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Симферополь
ИТ «АРИАЛ»
2018
УДК 504.73:57.085.2
ББК 28.59
М 67
Печатается по постановлению Ученого совета ФГБУН «Ордена Трудового Красного Знамени Никитский
ботанический сад – Национальный научный центр РАН» № 7 от «14» сентября 2018 г.
Оргкомитет конференции:
Председатель оргкомитета:
И.В. Митрофанова, д.б.н., заведующая отделом биологии развития растений, биотехнологии и
биобезопасности ФГБУН «Ордена Трудового Красного Знамени Никитский ботанический сад –
Национальный научный центр РАН».
Сопредседатели:
И.М. Донник, академик РАН, проф., д.б.н., вице-президент Российской академии наук;
В.А. Багиров, чл.-корр. РАН, д.б.н., директор Департамента координации деятельности организаций в
сфере сельскохозяйственных наук Министерства науки и высшего образования РФ;
Ю.В. Плугатарь, чл.-корр. РАН, д.с.-х.н., директор ФГБУН «НБС-ННЦ»;
О.И. Молканова, к.с.-х.н., заведующая лабораторией биотехнологии растений ФБГУН Главный
ботанический сад им. Н.В. Цицина РАН.
Члены международного научного комитета:
Dr. Jorge M. Canhoto, Professor, University of Coimbra, Department of Life Sciences, CEF - Centre for Functional
Ecology, Coimbra, Portugal; Dr. Maria Antonietta Germana, Associate Professor, Department of Agriculture,
Food and Forest Sciences (SAAF), University of Palermo, Palermo, Italу; Dr. S. Mohan Jain, Professor, Department
of Agricultural Sciences, University of Helsinki, Finland; Dr. Maurizio Lambardi, Professor, National Research
Council (CNR) of Italy, IVALSA/Trees and Timber Institute, Florence, Italy; Dr. Renato Paiva, Full Professor in
Plant Physiology, Department of Biology, Federal University of Lavras (UFLA), Lavras, Minas Gerais, Brazil;
Dr. Željko Prgomet, College Professor on Collegium Fluminense Polytechnic of Rijeka, Croatia; Dr. Kanchit
Thammasiri, Associate Professor, Department of Plant Science, Faculty of Science, Mahidol University, Bangkok,
Thailand; Dr. Alexsander Vainstein, Professor, The Robert H. Smith Institute of Plant Science and Genetics in
Agriculture, Faculty of Agriculture, Food and Environment, The Hebrew University of Jerusalem, Rehovot, Israel;
К.Ж. Жамбакин, чл.-корр АН РК, проф., д.б.н., генеральный директор РГП «Институт биологии и
биотехнологии растений» Комитета науки МОН Республики Казахстан; В.В. Титок, д.б.н., директор ГНУ
«Центральный ботанический сад Национальной академии наук Беларуси»; А.М. Кудрявцев, д.б.н.,
директора ФБГУН Институт общей генетики им. Н.И. Вавилова РАН; В.П. Упелниек, к.б.н., директор
ФБГУН Главный ботанический сад им. Н.В. Цицина РАН; П.Н. Харченко, академик РАН, научный
руководитель института ФГБНУ «Всероссийский научно-исследовательский институт
сельскохозяйственной биотехнологии»; О.В. Митрофанова, проф., д.б.н., главный научный сотрудник
лаб. биотехнологии и вирусологии растений ФГБУН «НБС-ННЦ»; И.Н. Третьякова, д.б.н., проф., ведущий
научный сотрудник ФГБУН Институт леса им. В.Н.Сукачева СО РАН; В.В. Корженевский, проф., д.б.н., зав.
лаб. флоры и растительности ФГБУН «НБС-ННЦ»; А.В. Смыков, д.б.н., зав. отд. плодовых культур ФГБУН
«НБС-ННЦ»; С.В. Шевченко, проф., д.б.н., главный научный сотрудник лаб. биохимии, физиологии и
репродуктивной биологии растений ФГБУН «НБС-ННЦ»; O.И. Коротков, к.с.-х.н. заместитель директора
по науке ФГБУН «НБС-ННЦ»; А.А. Веевник, к.б.н., заместитель директора по научной и инновационной
работе ГНУ «Центральный ботанический сад Национальной академии наук Беларуси»; И.Ю. Ковальчук,
к.с.-х.н. ведущий научный сотрудник лаб. криосохранения гермоплазмы Института биологии и
биотехнологии растений Комитета науки МОН Республики Казахстан; А.Е. Палий, к.б.н., заведующая лаб.
биохимии, физиологии и репродуктивной биологии растений ФГБУН «НБС-ННЦ»; Т.Б. Губанова, к.б.н.,
ведущий научный сотрудник лаб. биохимии, физиологии и репродуктивной биологии растений ФГБУН
«НБС-ННЦ».
Секретариат оргкомитета:
А.И. Ругузова, к.б.н. – секретарь; Н.Н. Иванова, к.б.н., старший научный сотрудник; В.А. Браилко,
научный сотрудник; С.В. Зубова, директор ООО «Научный сервис».
Члены рабочего оргкомитета:
Паштецкий А.В., Дуганов А.В., Лесникова-Седошенко Н.П., Челомбит С.В., Жданова И.В.,
Кузьмина Т.Н., Гребенникова О.В., Корзин В.В., Бакова Н.Н., Шишкин В.А., Андреев М.С.,
Овчинников С.А., Мельников В.А., Волик Л.Д.
Митрофанова И.В.
М 67 Материалы VIII Международной научно-практической конференции «Биотехнология как
инструмент сохранения биоразнообразия растительного мира (физиолого-биохимические,
эмбриологические, генетические и правовые аспекты)» ФГБУН «НБС-ННЦ», г. Ялта, Республика
Крым, Россия. 1 – 5 октября 2018 г. – Симферополь : ИТ «АРИАЛ», 2018. – 224 с.
ISBN 978-5-907118-13-3
УДК 504.73:57.085.2
ББК 28.59
© ФГБУН «НБС-ННЦ, 2018
ISBN 978-5-907118-13-3 © ИТ «АРИАЛ», 2018
4
Proceedings
of the VIII International Scientific and Practical Conference
«Biotechnology as an Instrument for Plant Biodiversity
Conservation (physiological, biochemical, embryological, genetic
and legal aspects)»
Yalta, Russia
October 01 - 05, 2018
Simferopol
РР «ARIAL»
2018
UDC 504.73:57.085.2
BBK 28.59
М 67
Published under the auspices by the International Society for Horticultural Science (ISHS)
Organizing Committee:
Chairperson: Irina Mitrofanova, Dr.Sci., the Head of Plant Developmental Biology, Biotechnology and Biosafety
Department, FSFIS «NBG-NSC».
Co-chairmen:
Irina Donnik, Academician of the RAS, Prof., Dr.Sci., Vice President of the RAS;
Vugar Bagirov, Corresponding Member of the RAS, Dr.Sci. the Director of Coordination and Providing Scientific
Organizations` Activities in Agricultural Sciences Department in the Ministry of Science and Higher Education of the
Russian Federation ;
Yurii Plugatar, Corresponding Member of the RAS, Dr.Sci., the Director of FSFIS «NBG-NSC»
Olga Molkanova, Ph.D., the Head of Plant Biotechnology Laboratory, FBSIS N.V. Tsytsin Main Botanical Garden of
the RAS.
International Scientific Organizing Committee:
Dr. Jorge M. Canhoto, Professor, University of Coimbra, Department of Life Sciences, CEF - Centre for Functional
Ecology, Coimbra, Portugal; Dr. Maria Antonietta Germana, Associate Professor, Department of Agriculture, Food
and Forest Sciences (SAAF), University of Palermo, Palermo, Italу; Dr. S. Mohan Jain, Professor, Department of
Agricultural Sciences, University of Helsinki, Finland; Dr. Maurizio Lambardi, Professor, National Research Council
(CNR) of Italy, IVALSA/Trees and Timber Institute, Florence, Italy; Dr. Renato Paiva, Full Professor in Plant
Physiology, Department of Biology, Federal University of Lavras (UFLA), Lavras, Minas Gerais, Brazil; Dr. Željko
Prgomet, College Professor on Collegium Fluminense Polytechnic of Rijeka, Croatia; Dr. Kanchit Thammasiri,
Associate Professor, Department of Plant Science, Faculty of Science, Mahidol University, Bangkok, Thailand; Dr.
Alexsander Vainstein, Professor, The Robert H. Smith Institute of Plant Science and Genetics in Agriculture, Faculty
of Agriculture, Food and Environment, The Hebrew University of Jerusalem, Rehovot, Israel; Kabyl Zhambakin,
Corresponding Member of the NAS RK, Prof., Dr.Sci., the Director of RSE «Institute of Plant Biology and
Biotechnology» of the Science Committee of the Ministry of Education and Science of Republic of Kazakhstan;
Vladimir Titok, Dr.Sci. the Director of the State Scientific Institution Central Botanical Garden of NAS of Belarus;
Alexander Kudryavtsev, Dr.Sci., the Director of FBSIS N.I. Vavilov Institute of General Genetics of RAS; Vladimir
Upelniek, PhD, the Director of N.V. Tsytsin Main Botanical Garden of RAS; Peter Kharchenko, Academician RAS,
Scintific Director of FSBSI “The All-Russian Research Institute for Agricultural Microbiology”; Olga Mitrofanova,
Dr.Sci., Prof. the Main Researcher of Plant Biotechnology and Virology Laboratory, FSFIS «NBG-NSC»; Iraida
Tretyakova, Dr.Sci., Prof., the Leading Researcher, Sukachev Institute of Forest SB RAS; Vladislav Korzhenevsky,
Dr.Sci. Prof. the Head of Flora and Vegetation Laboratory, FSFIS «NBG-NSC»; Anatoly Smykov, Dr.Sci, the Head of
Fruit Cultures Department, FSFIS «NBG-NSC»; Svetlana Shevchenko, Dr.Sci., Prof., the Main Researcher of Plant
Reproductive Biology and Physiology Laboratory, FSFIS «NBG-NSC»; Oleg Korotkov, PhD, Deputy Director for
Science, FSFIS «NBG-NSC»; Alexander Veevnik, Ph.D., Deputy Director for Scientific and Innovation Work “State
Scientific Institution Central Botanical Garden of NAS of Belarus”; Irina Kovalchuk, Ph.D., the Leading Researcher of
the Laboratory of Germplasm Cryopreservation at the Institute of Plant Biology and Biotechnology, Science
Committee of the Ministry of Education and Science of the Republic of Kazakhstan; Anfisa Paliy, Ph.D., the Head of
Plant Biochemistry, Physiology and Reproductive Biology Laboratory, FSFIS «NBG-NSC»; Tatyana Gubanova, Ph.D.,
the Leading Reseacher of Plant Biochemistry, Physiology and Reproductive Biology Laboratory, FSFIS «NBG-NSC».
Secretariat of the Organizing Commitee: Anna Ruguzova, Ph.D. – secretary; Natalya Ivanova, Ph.D., Senior
Researcher; Valentina Brailko, Researcher; Svetlana Zubova, the Director of “Scientific Service”.
Local Working Committee: Andrey Pashtetsky, PhD, Deputy Director; Anatoly Duganov, the Head of the
Department; Nina Lesnikova-Sedoshenko, Researcher; Svetlana Chelombit, Researcher; Irina Zhdanova,
Researcher; Tatyana Kuzmina, Ph.D., Senior Researcher; Oksana Grebenikova, Ph.D., Senior Researcher; Vadim
Korzin, Ph.D., Senior Researcher; Nadezhda Bakova, Ph.D., the Head of the Sector; Valery Shishkin, Ph.D., the Head
of the Laboratory; Michael Andreev, Research Engineer; Sergey Ovchinikov, Research Engineer; Vladimir
Melnikov, Researcher; Lylya Volik, the Head of the Department.
Mitrofanova I.V.
М 67 Proceedings of the of the VIII International Scientific and Practical Conference «Biotechnology as an Instrument
for Plant Biodiversity Conservation (physiological, biochemical, embryological, genetic and legal aspects)»
October, 1-5, 2018 – Simferopol : PP «ARIAL», 2018. – 224 p.
ISBN 978-5-907118-13-3
UDC 504.73:57.085.2
BBK 28.59
6
ОСНОВНЫЕ СПОНСОРЫ КОНФЕРЕНЦИИ
THE MAIN SPONSORS OF THE CONFERENCE
9
Welcome Address!
On behalf of the Organizing Committee, we are pleased to welcome all participants and
guests of the VIII scientific and practical conference "Biotechnology as an Instrument for
Plant Biodiversity Conservation (physiological, biochemical, embryological, genetic and legal
aspects)", which this year and for the first time in Russia is under the auspices by the
International Society for Horticultural Science (ISHS). International conference under this
name has become a traditional one and FSFIS "The Labour Red Banner Order Nikita
Botanical Gardens – National Scientific Center of the RAS", which has a valuable collection of
species and cultivars of decorative, subtropical and stone fruit crops, aromatic and medicinal
plants, takes this forum on its base for the third time.
Special attention is paid to the conservation of plant biodiversity in the world. The basis
of the conservation strategy of the species and cultivars of plants is determined by several
programming documents such as “Convention on Biological Diversity” (www.plant-
conservation–report-en.pdf), "Global Strategy for Plant Conservation" (www.botanicgardens/
ie/gspc/pdfs/gspc.pdf), "International Agenda for Botanic Gardens in Conservation"
(https://www.bgci.org/files/Worldwide/News/SeptDec12/international_agenda_web.pdf).
And biotechnology as an independent science on a new basis is intended to solve such
global problems as food, energy, resource security, environmental protection, etc. In many
countries the development of biotechnology is of great importance. Since the 80s of the past
century active development of new biotechnological methods has begun, and the end of the 20th
century and the beginning of the 21st century in the world were marked by the rapid
development of not only biotechnological researches, but also high-tech industries. Almost every
country in the world has its own biotechnology development program until 2020-2050s. In the
Nikitsky Botanical Gardens the works in the field of plant biotechnology were started 60-70s of
the past century. Due to comprehensive researches in the field of virology, phytopathology,
genetics, breeding, biochemistry, physiology and embryology of plants, it became possible to
actively develop and expand the scope of biotechnological researches. As a part of the program
of development of the scientific organization and the grant of the Russian Science Foundation
(2014-2018) № 14-50-00079 it has been managed to modernize the laboratory facilities of the
institution, creating new modern scientific complexes "BIOTRON" and "GENOMICS". The
construction of a new modern "PHYTOTRON" complex continues this work.
We gratefully acknowledge all key and invited speakers, participants with oral and
poster presentations for cooperation in the process of preparation and holding of the
conference.
We also want to express particular gratitude to the Federal Agency of Scientific
Organizations, all the sponsors of the conference, who tried to make the International
Biotechnology forum held at a high level.
With deepest respect and appreciation,
Irina Mitrofanova, Doctor of Biological Sciences,
Head of the Department of plant development biology, biotechnology and biosafety of FSFIS “NBG-NSC”,
Convener of “YaltaBiotech 2018” conference
10 10
ПЛЕНАРНЫЕ ДОКЛАДЫ
KEY NOTES
12
Haploid technology as valuable support to plant breeding in Citrus and
other fruit crops
Due to the characteristics of Citrus and of the majority of fruit crops, (such as the high
heterozygosity of the genomes, the long duration of the generation cycle, the large size,
and, often, the self-incompatibility), the traditional methods (involving several generations
of selfing), are not suitable to produce homozygous lines. The only feasible approach to
achieve homozygosity is “gametic embryogenesis”, that involving the regeneration from
gametes, products of meiotic segregation, allows the single-step development of complete
homozygous lines from heterozygous parents.
Haploids (Hs, plants with gametophytic chromosome number) and doubled haploids
(DHs, haploids that have undergone, spontaneously or in induced way, chromosome
duplication) are ever more goals of fruit breeding programs, as well as of many
agronomical important crops.
According to different genotypes, several innovative methods to produce Hs and DHs
are available, and among them, microspore embryogenesis through in vitro anther or
isolated microspore culture is one of the most efficient and extensively employed.
The purpose of this report is to provide some experiences, recent advancements and
forthcoming progresses on the application of DH technology as a powerful tool to support
plant breeding in fruit crops and especially in Citrus.
13
Genetic diversity, erosion, utilization and conservation under the
climate change
Global climate has continuously been changing, and is continuously changing gradually and
will have negative impact on global food production and food security in the coming future.
Recently erratic climate change, e.g. variation in annual rainfall, water deficit etc., has been
recorded worldwide that has affected food and agriculture. The developing countries have
more devastated impact due to poor infrastructure, lack of trained manpower and
economic conditions. Climate change and ever-growing human population are major
factors caused loss of genetic resources, arable land deletion and water shortage. Any small
change in global temperature may develop new insect and pests, and disease that may
devastate food and agriculture; and availability of sufficient water is a matter of concern to
sustainable agriculture. Genetic diversity is the key for the survival, evolution of species
and utilization for crop improvement. Genetic variation within a species is important for its
ability to adapt to a changing environment. Species having larger levels of genetic diversity
have a better chance of adaptation, survival and deployment over a wide range of
environmental conditions. Appropriate levels of genetic variation should be maintained in
the populations of a species for conservation planning and prevent genetic erosion. The
conservation of genetic resources should be based on the genetic architecture and
phenology, and how genetic and phenotypic variation is organized and distributed within
and among populations of a species. Induced genetic diversity is caused by radiation and
chemical mutagens in most of the major crops, and used for developing new varieties with
useful traits that had great economic benefit to growers and food supply chain. However,
climate change is a greater challenge on crop production, e.g. water availability, soil quality,
insect and pests. Plant genetic diversity is conserved by cryopreservation (somatic
embryos, embryogenic cell cultures), cold storage (seed and in vitro shoots) and in vivo
(field gene banks), establish germplasm website for exchange and utilization. Plant
regeneration from somatic embryos and embryogenic cell suspension is necessary for
applying cryopreservation by using liquid nitrogen. In cold storage, shoot cultures are
preserved at 4-5°C; however, subcultures are needed even though their number is reduced.
In cryopreservation, subculture is not needed and cultures are stored for longer period.
Seed banks are commonly used in most of the seed crops. Field gene bank is alternate to in
vitro conservation, and is being widely used, however has risk of insect and pest attack. We
will discuss the importance of genetic diversity conservation, climate change and setting up
of gene banks.
Keywords: plant genetic diversity, cold storage, cryopreservation, seed banks, in vitro
conservation, climate change.
14 14
Biotechnology for plant genetic resources conservation: an overview of
in vitro-banking and cryobanking in the world
Today, in vitro culture is a strategic tool to support medium and long-term conservation of
plant genetic resources by using the slow growth storage of shoot cultures and the
cryopreservation of organs and tissues. Over the last 30 years, considerable progresses
were made in the development of both techniques that are nowadays considered as ex situ
conservation strategies complementary to traditional seed banks and in-field clonal
collections. Conservation in slow growth storage consists modifying the medium and/or
culture conditions to reduce the growth of in vitro shoots without affecting their viability
and regrowth potential when moved back to standard culture conditions. The technique
allows medium-term conservation of shoot cultures from a few months to two years and
more. Cryopreservation preserves plant organs and tissues at the ultra-low temperature of
liquid nitrogen (-196°C).
Currently, almost 45,000 accessions of vegetables (mainly potato, cassava, sweet
potato, yam and garlic) and fruit species (mainly apple, mulberry and banana) are
maintained in 21 genetic resources conservation centers, located in 16 Countries and 5
Continents (Europe, Asia, Africa, North and South America). Over 70% of these accessions
are maintained in vitro by means of slow growth storage of shoot cultures, but more recent
cryopreservation is constantly growing, and over 12,500 accessions from vegetables are
stored at -196 °C. In order to avoid too many replications of the same genetic resources in
the different operating centers, a networking work was recently undertaken under the
aegis of Bioversity International, the Global Crop Diversity Trust and the International
Potato Center and led, in July 2017, to a report containing a census on plant genetic
resources in in vitro-banks and in cryobanks, as well as indications and directives on the
organization and costs of these forms of conservation. A first important step towards a
global coordination of conservation strategies in vitro and in liquid nitrogen which will
soon become indispensable.
15
Somatic embryogenesis and organogenesis as biotechnological ways of
some horticultural plant propagation and conservation
Irina Mitrofanova
Federal State Funded Institution of Science "The Labor Red Banner Order Nikita Botanical
Gardens – National Scientific Center of the RAS", Yalta, 298648, Russian Federation;
E-mail: irimitrofanova@yandex.ru
Studies on the morphogenesis in vitro in some ornamental, essential oil and subtropical fruit
crops, as well as the development of preservation protocols, have been carried out by the
support of the Russian Science Foundation (research grant No. 14-50-00079).
Keywords: fruit, ornamental and essential oil plants, direct and indirect regeneration,
slow-growing plant collection, in vitro.
16 16
Соматический эмбриогенез и органогенез как биотехнологические
пути размножения и сохранения некоторых садовых растений
И.В. Митрофанова
ФГБУН «Ордена Трудового Красного Знамени Никитский ботанический сад –
Национальный научный центр РАН», Никитский спуск 52, пгт Никита, г. Ялта,
298648, Российская Федерация; e-mail: irimitrofanova@yandex.ru
17
Development of cryopreservation of plant germplasm
Kanchit Thammasiri
Department of Plant Science, Faculty of Science, Mahidol University, Rama VI Road,
Phyathai, Bangkok 10400, Thailand; E-mail: kanchitthammasiri@gmail.com
Plant cryopreservation started by Prof. Dr. Akira Sakai about 60 years ago. The methods
were developed rapidly by many scientists from dormant buds to slow freezing,
encapsulation-dehydration, vitrification, encapsulation-vitrification, droplet-vitrification,
V-cryo-plate and D-cryo-plate with different types of dehydration, respectively. The
knowledge and idea came from the notice of season change and plant survival in cold
weather. The two principles for plant cryopreservation methods are: 1. slow freezing and 2.
vitrification. At present, various methods were applied for many plant species in many
genebanks around the world. This presentation will discuss each plant cryopreservation
method and shows some methods that were applied at Mahidol University, as well as
discussing the future of plant cryopreservation.
18 18
Specialized metabolic pathways: cues controlling floral scent and color
production
Alexander Vainstein
The Hebrew University of Jerusalem, POBox 12, Rehovot, Israel;
E-mail: alexander.vainstein@mail.huji.ac.il
The survival of many plant species depends on their ability to attract pollinators.
Exhibitionistic characters, such as flower color and fragrance, determined largely by
specialized metabolites, are among the main means adopted by plants to achieve this goal.
Using virus-induced gene silencing/editing and expression approaches for large-scale
identification of floral scent genes, we identified and characterized petunia flower-specific
scent-regulatory genes named EMISSION OF BENZENOIDS (EOBs). We further
demonstrated that PH4, a myb gene that determines final petal hue through pH
homeostasis in the early stages of flower development, is essential for emission, but not
production, of the floral volatile bouquet in mature flowers. We also revealed that petunia
flowers produce significant amounts of sugar-conjugated scent compounds with a unique
diel accumulation pattern that are further catabolized in parallel to scent emission. These
phenylpropanoid glycosides are stored in the vacuoles of petunia flowers, providing first
evidence of subcellular compartmentalization of scent compounds.
We revealed that gibberellin (GA) acts as a developmental cue regulating floral scent
production in petunia. GA-dependent timing of the sequential activation of different
branches of the phenylpropanoid pathway, products of which are responsible for either
color or scent—both necessary for pollinator attraction—may represent an intriguing
machinery developed by plants to enable intimate crosstalk between floral pollination
syndromes.
Knowledge gained from an understanding of the molecular mechanisms underlying
the phenomenon of floral fragrance/pigmentation will enable better insight into nature's
way of ensuring evolutionary success and should also provide general ideas on in-planta
networking for secondary metabolism allowing in the future rational design of plants’
color/scent pallet.
19
20
УСТНЫЕ ДОКЛАДЫ
ORAL PRESENTATIONS
22
Clonal micropropagation of some Jerusalem artichoke cultivars and
molecular characterization using RAPD and ISSR based markers
23
The diversity of fruit shapes of Prunus armeniaca L. natural populations
in Mountainous Dagestan
The study of natural apricot populations in Dagestan has a great significance for solving
series of questions about taxonomy, genetics, ecology and breeding. Comparative analysis
of Prunus armeniaca L. 418 trees from Mountainous Dagestan natural populations on the
qualitative characteristics of the fruit showed that 91% have small-sized fruits with a
rounded shape (77%), a flat top (71%) with predominance of orange skin color (69%) and
pulp (70%) with acidic taste (74%) and dry consistency (56%). Some kinds of tree (up to
26%) has fruits with a hard and partially detached flesh from the stone, with a blush
(40%). It is not found practically large-fruited forms, as well as forms with cream-colored
fruit and flesh. It is found out that the phenotypic variety of fruits decreases, the percentage
of rounded dried fruit small-bodied with orange coloring of the fruit with acidic taste and
hard-to-remove stone increases according to the distance from the gardens and increasing
of the height above sea level. The clustering of natural apricot populations in Dagestan has
shown that "fetal shape" is the main differentiating sign. The significant role in the
grouping of apricot has the signs of "flesh color" and "skin color", but "fruit taste", "top
form" and "fruit consistency" has a lesser extent. The signs "form of foundation", "depth of
the seam", "presence of blush" and "separation from the stone" are not very informative.
24 24
Формовое разнообразие плодов природных популяций
Prunus armeniaca L. Горного Дагестана
25
Dynamics of antioxidants accumulation in tea (Camellia sinensis (L.)
Kuntze) at the conditions of Russia subtropics
Many plants contain substances with antioxidant activity. These substances include
antioxidant enzymes (for example, peroxidase, catalase, SOD, etc.) and non-enzymatic
antioxidants (ascorbic acid, carotenoids, flavonoids, etc.). It is believed that antioxidants,
changing in metabolic reactions the dynamics of many enzymes and possessing
antibacterial action, are indispensable components of the products for humans. This quality
is from tea plants. Tea, as a product included in the daily diet of a person, is the richest in
antioxidants. One of the strong tea antioxidants is ascorbic acid (AA), it plays an important
role in the regulation of redox processes. AA is closely related to plant phenol from the
group of flavonols - vitamin P (rutin). Rutin increases the activity of ascorbic acid,
protecting it from oxidation. Objects of study were samples of black and green tea made
from 2-3 leaf flushes of ‘Kolchida’ and ‘Sochi’ cultivars, radiated mutant № 3823 and №
582, chemical mutant № 855, colchicum mutant № 2264, grown for an experienced
collection area, founded in 1984 – 1985 at the village of Uch-Dere (Sochi, Lazarevsky
region). We show dynamics of accumulation of vitamins C and P in black and green tea
during months. Accumulation of vitamin P (1.5 – 1.7 times) and AA (2.7 – 3.2 times) from
May to August were increased, and in green tea the rutin content in 3 – 4 times, and AA
almost in 4 – 5 times higher than in black. In the accumulation of vitamins C and P of
varietal characteristics are occured. For green tea, the highest number of AA was observed
in samples produced from flushes of cultivar ‘Sochi’ and radiated mutant № 3823 (48.03
and 44.94 mg%, respectively), and the lowest in colchicum mutant № 2264 (29.60 mg%).
In black tea, AA contains radiated mutant № 3828, chemical mutant № 855 and cultivar
‘Sochi’ (9.53; 9.28 and 9.47 mg%, respectively). Other distribution is revealed in contents of
vitamin P, in the samples containing more quantity of AA it is noted less rutin, than in tea
with the low contents of AA.
Keywords: tea, cultivar, mutant form, antioxidant, ascorbic acid, vitamin P, dynamics.
26 26
Динамика накопления антиоксидантов в чае (Camellia sinensis (L.)
Kuntze) в условиях субтропиков России
27
The creation of modern gene pool banks, a fundamental task for solving
the import substitution problem
In accordance with the Decree of the President of the Russian Federation of July 21, 2016
No. 350 "On Measures for the Implementation of the State Scientific and Technical Policy in
the Interests of Agricultural Development", as well as Decree No. 996 of the Government of
the Russian Federation of August 25, 2017 "On Approval of the Federal scientific and
technical program for the development of agriculture for 2017-2025", conservation and
replenishment of biodiversity in Russia is a priority task for import substitution. The
creation of gene pool banks of fruit and small fruit plants that must be laid in accordance
with international norms and standards should be based on planting material free from
harmful viruses and dangerous pathogens, which are typical plants tested for productivity
and genetic stability. Along with the formation of field collections, the use of
cryopreservation in vitro of healthy specimens is envisaged. With such system, the task of
creating and introducing new cultivars of fruit and small fruit crops into industrial
plantations in Russia, which are now often laid by imported planting stock, will be
accelerated by orders of magnitude for the established breeding and nursery centers.
Possessing original plants of promising cultivars and hybrids or their clones (if
biotechnological methods and cryopreservation were used in the breeding process),
nurserymens, having patented domestic developments, will be able to reproduce them in
the shortest terms in conditions of modern protected soil without violating genetic
stability. In complex studies conducted by breeders, biotechnologists, virologists,
phytosanitary experts and nurseries, not only digital technologies will be born, but also
mother plantings of the highest quality categories. Only in this way the scientifically-based
system of nursery management, elaborated by the scientists of the All-Russian
Horticultural Institute for Breeding, Agrotechnology and Nursery (ARHIBAN), will work.
The role of the state here is paramount, since it is necessary to provide comfortable
conditions for conducting scientific research and efficient business functioning.
Today, ARHIBAN has all the prerequisites for the implementation of its developments
in such conditions, supporting innovative technologies by transferring planting material of
higher quality categories for the laying of mother plantations in accordance with the
scientifically based system of nursery management in Russia.
Keywords: import substitution, gene-pool banks, breeding and nursery centers, source
plants, digital technologies.
28 28
Создание современных генбанков – фундаментальная задача для
решения проблемы импортозамещения
29
Determination of drought resistance of common fig and jujube cultivars
through leaf dehydration
Investigation of fruit-bearing crops efficiency proves that a yield quantity and quality
directly depend on plant stages and susceptibility to the environmental factors. Lately in
many regions, including the Crimea, the specialists have fixed changes of the temperature
regimes. The number of extremely hot days (over 31°C) considerably have grown. In case
of a high temperature and moisture deficit, leaves of common fig and jujube started wilting
earlier than ever, lost turgor, slowed down growth and decreased fruiting. At the same
time, moisture deficit in the organs and tissue causes crop capacity reduction and
aggravation of a fruit quality. Fruits don`t gain the size and taste parameters, which are
typical for a certain cultivar. Purpose of this work is to study characteristics of moisture
regime and potential draught-resistance of different cultivars and forms of Ziziphus
jujuba Mill. and Ficus carica L. under the conditions of the Southern Coast of the Crimea.
The researches were being carried out during 2015-2017 based on the collection
plantations of the Nikita Botanical Garden (NBG). The samples of domestic and foreign
breeding were taken as the study objects. While rating a draught-resistance, capacity of
water-retaining and turgor regenerating in leaf tissue were determined after a wilting
process during maximum high air temperature and aridity. There is a direct dependence
between a leaf capacity to retain moisture and degree of a turgor regenerating, that makes
possible to evaluate a plant potential in resistance to dehydrating factors. If long-term
dehydrating (30 hours), the most of the jujube cultivars and forms from NBG breeding
presented a high reparation capacity - from 70 to 90%. The following jujube cultivars and
forms were distinguished with a high draught-resistance: ‘Tsukerkovy’, ‘Koktebel’, ‘Pl. 5-
27’. The close relation between a water-retaining capacity and leaf adoptability to stand by
the dehydration was determined as the result. Wilting leaves output moisture slowly as
possess resistance to dehydration. The cultivars and forms of common fig in ‘Sabrutsija
Rozovaja’, ‘Figue Blanche’ and ‘Figue Jaune’ were distinguished according to this
parameter. Turgor retaining in leaves tissues ranged from 70 to 100%. The cultivars and
forms with a high water-retaining capacity during critical points of vegetation can be
recommended for a breeding work and creating a new plantation in extra arid regions.
This study was funded by a research grant No. 14-50-00079 of the Russian Science
Foundation.
30 30
Определение засухоустойчивости сортов инжира и зизифуса по
обезвоживанию листьев
31
The growth characteristics of Abies alba seedlings in Voronezh
Artamonova str. 8a, 33, 394010, Voronezh, Russian Federation; E-mail: olya34@mail.ru
This research focuses on growth and development of seedlings of European silver fir
(Abies albaL.), native to the Central Europe, in the Central Black Earth Region. The research
aims at introducing this species to forestry and green building in the region. The study is
based on measurements and observations of silver fir seedlings in the forest nursery of All-
Russian Research Institute of Forest Genetics, Breeding and Biotechnology, Voronezh. The
study started in 2010. We used silver fir from the arboretum of Voronezh State University
of Forestry and Technologies as parent trees, which were planted in 1992 by five-year
seedlings. They first produced seeds in 2009, at the age of 22 years. In 2010 we collected
seeds and sowed them after pre-sowing seed treatment. Their germination capacity was
about 90%. After 10-14 days the shoots appeared. By the end of the first growing season
the height of a seedlings was 7.2 cm on average. We continued to monitor their growth and
physiological status on an annual basis. The study demonstrates that two key determinants
of plant growth are temperature and water availability, and during the first half of the
growing season, when soil still has enough water, plant growth depends primarily on the
sum of positive temperatures while during the second half of it, due to water shortage,
upon precipitation patterns. During the final year of the research the average fir growth
was significantly higher than before. The inter-series correlation coefficient between the
years was nearly the same. The first-order autocorrelation between the growth of the
current and previous year was 20-25%, which indicates a stable growth and development
of the seedlings.
Thus, it can be concluded that the silver fir is quite adapted to the conditions of the
region and can be recommended for introduction into forestry practice.
32 32
Особенности роста сеянцев Abies alba в условиях Воронежа
33
Ultrasound diagnostics of some species of Cupresus L. genus in the plant
collection of the NBG-NSC
The studies have been performed in situ on the territory of the parks of Arboretum of the
Nikita Botanical Gardens. «Arbotom®» a complex of acoustic ultrasound tomography has
been used to investigate the state of trunk wood tissue of Cupressus species, which are
widely used in landscape design in the Southern Coast of the Crimea (SCC): Cupressus
torulosa D.Don, Cupressus arizonica Greene, Cupressus sempervirens L. (var. stricta Ait) and
Cupressus macrocarpa Hartw. The low rate of phytogenic damage is observed in C. torulosa.
The trunk wood tissues of C. marcocarpa are characterized by the highest rates of
phytopathogenic degradation. The graphic presentation of ultrasound scanning in most
part of trunk wood volume of C. macrocarpa is defined by red and purple colors of marking,
which indicates the last stage of trunk wood tissues degradation and the dramatical
decrease of vegetal functions of damaged plants. Correlation analyses of phytopathogenic
damage’s rate and dendrometric features of the studied species of Cupressus genus has
shown, that the intensity of development of root rot at stumpwood has a positive relation
with its sizes. It has been revealed that the species in vegetative phase are the most
vulnerable to the adverse effect of pore fungi. The correlation between the rate of
phytopathogenic damage of trunk wood and the external indexes of vital state is observed
at a tendency level. It is determined by the fact that the external symptoms of the disease
affection do not appear at once. In many cases, the visualization of damages is noticed at
the stage of phytopathogen’s fruiting body formation, when the destructive processes
spread over the most part of trunk wood. The negative impact of pore fungi is one of the
most significant factors decreasing the vegetative functions and limiting the longevity
of Cupressus genus’ species under the conditions of the SCC. The phytopathogenic damage
of wood and the formation of stem rot have an effect on the implementation of the bio-
environmental program of growth and development of a plant, reducing the time scale of
passing certain ontogenesis stages.
This study was funded by a research grant No. 14-50-00079 of the Russian Science
Foundation.
Keywords: Cupressus L., wood tissues, phytopathogenic damage, tomography, vital state.
34 34
Ультразвуковая диагностика состояния растений некоторых видов
рода Cupressus L. коллекционных насаждений НБС-ННЦ
35
Water regime peculiarities of apricot cultivars and breeding forms in
conditions of the Southern Coast of Crimea
Adaptation to environmental conditions is one of the main reasons for apricot plants that
limit their spreading to different climatic zones. A great variety of soil-climatic conditions
of natural regions determines the amplitude of the requirements for cultivars. The
breeders are focused on the degree of adaptation of the genotype being created.
The aim of the research was to study the drought resistance of apricot cultivars and
breeding forms in conditions of the Southern Coast of Crimea, to identify the degree of their
adaptation to the conditions of cultivation and to select promising samples for further use
in breeding for this trait. The 18 genotypes of plants have been studied during 2015-2017.
Water-cut of leaf tissues, water retention capacity and restoration of turgor were
determined by the methods of G. Eremeev (1974); water deficit was estimated according to
the methodology of M. Kushnirenko (1975). It was pointed out that the cultivars and
breeding forms of the Nikita Botanical Gardens are more adapted to the droughty
conditions of the southern region of Russia than the introduced species. The influence of
meteorological conditions of the year on the water-cut of the leaves of apricot is noted. Its
greatest variation was recorded in 2015 (from 23.1 to 67.9%, depending on the genotype).
This year, the average monthly air temperature has been exceeded by 1.2-2.9°C compared
to average long-term standards and the amount of precipitation decreased by 11.6-15.4
mm. In the course of the experiment, it was established that the loss of water during the
withering of leaves depends on the characteristics of the cultivar. The greatest amount of
moisture after 24 hours of wilting following the results of 2015-2017 preserved three
plants. In three years of study, there were eight selected promising cultivars and breeding
forms by the set of indicators.
This study was funded by a research grant № 14-50-00079 of the Russian Science Foundation.
Keywords: apricot, cultivars, selection forms, drought resistance, water-cut, water deficit,
turgor restoration.
36 36
Особенности водного режима сортов и селекционных форм
абрикоса в условиях Южного Берега Крыма
37
Genetic diversity assessment of Crimean wild grape forms based on
microsatellites polymorphism
The Crimean peninsula due to its favorable environmental and geological conditions can be
considered as an area for survival and development of Vitis vinifera subsp. sylvestris
(Gmel.). It is believed that wild Vitis vinifera L. vines, including V. vinifera ssp. sativa D.C.
that fell out of cultivation and true wild grape V. vinifera ssp. silvestris Gmel. that survived
since the tertiary period and is believed to be the progenitor of Vitis vinifera, can be found
in the Crimean mountains. To identify habitats of wild growing grapes, the forested areas of
the Crimean mountain-forest natural reserves were examined in the period of 2015-2018.
The territory of Eski-Kermen and Mangup cave towns, the slopes of Baydarskaya,
Yaltinskaya and Alushtinskaya yailas, the gorge of the Chernorechensky canyon and the
couloirs along Alaka, Kalenda, Armanka and others rivers were examined. A total of 345
wild-growing lianas were detected. The identified vine plants primarily grew at 600 m
above sea level in areas with sufficiently moisturized soil. Image-documentation of a shoot,
leaf, inflorescence and cluster (when available) was done. Molecular assessment of the
grape genotypes was performed based on analysis of allelic polymorphism of nine nuclear
microsatellite loci. The standard genetic parameters were calculated using the POPGENE
software (v.1.32); the DARwin software (v.5.0.158) was used to construct a phylogenetic
tree. The share of polymorphic loci made 100%. The average number of alleles/loci was
8.56, the effective number of alleles (ne) was 3.6, the Shannon information index (I) made
1.57. A large number of rare alleles was recorded (25.4%). In the wild vine samples, a
heterozygote deficiency was revealed – the average value of the Wright fixation index
(coefficient of inbreeding) Fis = 0.107.
The research is conducted with the support of the Russian Foundation of Basic Research,
grant No. 15-29-02715.
Keywords: Vitis vinifera ssp. silvestris Gmel., SSR, allelic polymorphism, genetic variability.
38 38
Оценка генетического разнообразия крымских дикорастущих форм
винограда на основе полиморфизма микросателлитных локусов
С.М. Гориславец, В.А. Володин, Я.А. Волков, А.А. Колосова, Е.П. Странишевская,
В.И. Рисованная
Федеральное государственное бюджетное учреждение науки «Всероссийский
национальный научно-исследовательский институт виноградарства и виноделия
«Магарач» РАН», ул. Кирова, 31, г. Ялта, 298600, Российская Федерация;
e-mail: MGR.Magarach@gmail.com
Ключевые слова: Vitis vinifera spp silvestris Gmel., SSR, аллельный полиморфизм,
генетическая изменчивость.
39
Old abandoned terraces surveying and restoration as a contribution to
the adaptive forest landscape restoration of the Shouf Biosphere
Reserve (Lebanon)
Nizar Hani1, Pedro Regato2, Marco Pagliani2, Mishal Khaddazh3, Yuri Vernyuk3, Rawya Bu
Hussein1, Lina Sarkis4, Monxer Buwadi1, Vadim Plyushchikov3
1Shouf Biosphere Reserve (SBR), Park House, Village Square,Maaser, Lebanon;
2SBR International consultants, San Gerardo 8, 28035 Madrid, Spain;
3Peoples’ Friendship University of Russia (RUDN University), Miklukho-Maklaya, 6,
The Shouf Biosphere Reserve (SBR), with a surface of 50,000 hectares is located in Mount
Lebanon. SBR harbors a rich flora with about 1,100 species, many of them with edible,
aromatic and medicinal properties. The landscape includes a mosaic of agro-silvo-pastoral
systems, characterized by pastures, forests - including the largest expanse of Cedrus libani
forests in Lebanon – and traditional agriculture terraces (olives, fruit trees). Landscape in
the SBR is the result of: (i) overuse of habitats and wild plant species; (ii) inadequate
policies on land use planning; (iii) rural abandonment resulting in soil erosion, agro-
biodiversity loss, and the conversion of agricultural land into chaotic urbanization; (iv)
agriculture intensification, pollution. Climate change trends in the region are already
exacerbating environmental risks (forest fires, water scarcity). Building resilience in the
SBR through forest landscape restoration entails interventions to strengthen the adaptive
capacity of the social and ecological politics in the region. The project developed protocols
for the production and sustainable use of high quality plant material of more than 40 native
plant species. An advanced method for monitoring and large scale detailed mapping was
used to improve the investigation of terraces. More than 12 hectares of degraded terraces
were restored and planted with native fruit tree species and shrubs. Sustainable farming in
the restored terraces implied combined actions on: (i) biodiversity conservation – both the
diversification of planted species and cultivars, and the conservation and management of
marginal habitats linked to farming system; (ii) organic food production, and (iii) the
management of agriculture waste to reduce environmental risks while generating new
revenues.
40 40
Осмотр и восстановление старых заброшенных террас в качестве
вклада в восстановление адаптивного лесного ландшафта
биосферного заповедника Шуф (Ливан)
Низар Хани1, Педро Регато2, Марко Паглиани2, Мишаль Хаддаж3, Ю.И. Вернюк3,
Рауья Бу Хусейн1, Лина Саркис4, Монзер Бувади1; В.Г. Плющиков3
1Биосферный заповедник Шуф (БЗШ), г. Маасер, Ливан;
2Международные консультанты (БЗШ) , ул. Сан Герардо, 8, г. Мадрид, 28035,
Испания;
3Российский университет дружбы народов (РУДН), ул. Миклухо-Маклая, 6, г. Москва,
41
Effect of some modifications in tissue culture media for commercial
banana multiplication
Giza, Egypt
The goal of this study was to find cheaper alternatives for micropropagation of banana
(Musa sp.). Low-cost options adopted in this study included lowering MS concentration in
media, lowering benzylaminopurine (BAP) in media, and replacement of distilled water
with tap water in media preparation. The effect of 4 different concentrations of BAP (0.6
mg L-1, 0.8 mg L-1, 1.0 mg L-1, 1.2 mg L-1) on bud development initiation and shoot length
was investigated. The experiment tested the effect of lowering MS concentration in media
with threedifferent concentrations (3.0 g L-1, 3.5 g L-1, 4.4 g L-1) in order to reduce the cost
of media ingredients. The effect of replacing distilled water with tap water was also
studied. The investigated concentrations of BAP has shown that using lower concentrations
of BAP (0.6 mg L-1, 0.8 mg L-1) will produce a slightly lower number of buds while
producing higher shoot length if compared to standard concentrations (1.0 mg L-1, 1.2
mg L-1). The study shows that using low MS concentration (3.0 g L-1) provides a higher
number of bud development and shoot length when compared to the standard (4.4 g L-1).
Using tap water instead of distilled water was effective in lowering cost of production
without compromising the quality of plants. Tested media with lowered ingredients (BAP:
0.6 mg L-1, MS: 3.0 g L-1, tap water) was tested against standard media and it showed no
difference in the number of bud dvelopment while producing higher shoot length plantlets.
Using the tested media will cut the cost of media preparation by nearly 50%.
42 42
The use of high-throughput sequencing to investigate the diversity of
defense protein genes of cucumber inoculated with cucumber green
mottle mosaic virus
Cucumber green mottle mosaic virus (CGMMV) is an RNA virus from tobamoviruses group
that infects plants from the Cucurbitaceae family, causing a loss of up to 40% of the crop.
The resistance genes to CGMMV have not been described, so attenuated strains are used for
plant protection, which do not cause symptoms of the disease, but provide protection
against pathogenic strains. CGMMV genome regions determined its pathogenicity were
revealed earlier, but no studies were conducted to investigate the changes in transcriptome
of cucumber Cucumis sativus upon its inoculation with viruses, which allow detecting
proteins forming plant immune response, including antimicrobial peptides (AMP). Here
this investigation was made by sequencing the transcriptomes of eight cucumber samples
in two technical repeats on Illumina Hiseq 2500 platform: healthy plants, plants treated
with an attenuated and pathogenic strain, and vaccinated plants to which an additional
pathogenic virus strain was applied. About 50 million reads with a length of 150 bp were
obtained for each sample. Transcriptome assembly was performed using Trinity. The
number of transcripts obtained by the program was 55,000-65,000 with the median
transcript length in the range of 840-961. The search for plant defense proteins in the
transcripts obtained using Blast2Go, SPADA and Cysmotif searcher allowed to reveal about
100 proteins involved in response to pathogens in each of the samples. In addition, infected
and vaccinated plants had more AMPs than healthy plants. Preliminary data on defense
proteins differential expression allow identifying transcripts involved in the genetic
regulation of the plant response to inoculation by various strains of CGMMV. Currently we
perform additional investigations of the regulation and searching for new AMPs, and to
develop practical applications of new defense proteins revealed in the process of
sequencing data analysis.
This work was supported by the Russian Science Foundation (Project No. 16-16-00032).
43
Использование высокопроизводительного секвенирования для
исследования разнообразия генов защитных белков огурца, при
заражении вирусом зеленой крапчатой мозаики огурца
44 44
Effect of cold shock proteins on increasing cold and frost resistance in
winter rape
Chemistry of the RAS, Prospect Nauki, 6, Puschino, Moscow region, 142290, Russian
Federation;
2 All-Russia Research Institute of Agricultural Biotechnology, Timirayzevskaya Str., 42,
Gardens – National Scientific Center of the RAS", Yalta, 298648, Russian Federation;
E-mail: dolgov@bibch.ru
The increased demand for vegetable oil for food purposes and high - protein feed for
livestock and poultry encourages Russian producers to expand the production of various
oil crops, while occupying as the acreage of the southern part of Russian central region and
Volga region, where the winter frosts with the temperatures below -20°C are common.
Improvement of cold and frost resistance of cultivated crops would significantly increase
yield and the profitability of oil production and expand the range of rape cultivation in
Russia. Domestically selected winter rape of the ‘Severyanin’ cultivar was used as an object
of research. Commercial value of this cultivar is high seed and fodder productivity, high
quality of oil and meal, and resistance to major diseases. The vector constructs pBI121-
CSPA-plant and pBI-TsCSDP3, which were created on the basis of pBI121 vector by
replacing the ß-glucoronidase synthesis gene with the target cspA gene isolated from
Escherichia coli, with the optimized codon composition for the plants of the cabbage family
(Brassicaceae) (cspA-plant), and the target tscsdp3 gene, encoding the protein with the cold
shock domain of Thellungiella salsuginea (Pall.), were used in the experiments,
respectively. Transformation was performed by co-cultivation of hypocotyl explants with
Agrobacteria for 48 hours. As a result of the studies, 9 kanamycin-resistant lines were
obtained. PCR analysis revealed 6 transgenic rapeseed lines with the introduction of the T-
DNA construct pBI121-CSPA-plant and 3 lines with the insertion of the T-region of plasmid
pBI-TsCSDP3.These lines were adapted to in vivo conditions and planted in a greenhouse
to obtain the seed generation T1. After germination of T1 generation seeds in the presence
of 650 mg L-1 kanamycin, 40 resistant plants of each line were selected for assays to
determine the expression level of heterologous genes and to identify the effectiveness of
introduced genes for increasing the cold resistance of transgenic rapeseed lines.
Studies were carried out in accordance with the State Order (0574-2014-0024), State
registration number АААА-А17-117082950004-9 and State Order (0829-2016-0001), State
registration АААА-А18-118012300084-8.
45
Изучение эффективности применения белков холодового шока для
повышения холодостойкости и морозоустойчивости озимого рапса
Национальный научный центр РАН», Никитский спуск 52, пгт Никита, г. Ялта,
298648, Российская Федерация; E-mail: dolgov@bibch.ru
46 46
Evaluation of the Nikita Botanical Gardens genefund collection of
persimmon with parameters of the ideal cultivar
Persimmon Oriental, as a fruit crop, has become widespread almost on all continents of our
planet. In the fruit growing of the Crimea, persimmon has been actively cultivated since the
beginning of the XIX century. However, in practice, introduced cultivars of foreign breeding
acclimatize in the soil and climatic conditions of the Crimean peninsula with great
difficulty. To solve this problem, the Department of Subtropical Fruit Crops was established
in the Nikita Botanical Gardens in 1937, whose staff began a complex works on the
introduction and breeding of persimmons using scientifically based approaches (Clyde,
1948). The approaches and methods used in the breeding of fruit crops are continuously
improved. For complex evaluation of promising and classic cultivars, the model of an ideal
persimmon cultivar was developed based on the following characteristics: yield (20 t/ha),
weak affection by fungal diseases (1 point), fruit mass (200 g), lack of fruity stiffness in the
solid state (3 points ), sugar content (25%), fruit taste (5 points), absence of seeds (3
points). For comparison with the model, 13 cultivars of persimmon were selected from the
genefund collection of the Nikita Botanical Gardens. The studies results were processed
using the Statistics 10 program by the cluster analysis method. As a result, persimmon
cultivars with economically valuable characteriestics corresponded to the parameters of
the model of the ideal cultivar have been selected. By the similarity and magnitude of the
common characteristics persimmons are grouped into clusters (groups): I – ‘Zolotistaya’; II
– ‘Hyakume’, ‘Hachia’, ‘Nikitskaya Bordovaya’, ‘Yuzhnaya Krasavitsa’, ‘Mechta’, ‘Sidles’,
‘Gayley’; III – ‘Costata’, ‘Zvezdochka’, ‘Zenjimaru’, ‘Fuyu’, ‘Nikitsky Prevoskhodny’.
According to the results of the complex evaluation, the closest to the ideal model are the
following cultivars: ‘Zolotistaya’, ‘Costata’, ‘Zvezdochka’, ‘Mechta’ and ‘Sidles’.
This study was funded by a research grant № 14-5-00079 of the Russian Science Foundation.
47
Оценка генофондовой коллекции хурмы Никитского ботанического
сада по параметрам идеального сорта
48 48
Somatic hybridisation as a potential tool for Pelargonium improvement
Every year, millions of garden geraniums are being produced for decoration throughout
the world. They have been derived from the wild species of Pelargonium indigenous to
South Africa and crossings between them. Interspecific crossings may allow the use of the
broad genetic pool of the genus Pelargonium. As such crossings remain difficulty, somatic
hybridisation could be an alternative to overcome crossing barriers. Here, we present the
production of asymmetric somatic hybrids between P. × hortorum ‘Antik Pink’ and
Pelargonium wild type 662. Hypocotyl protoplasts of ‘Antik Pink’ and mesophyll
protoplasts of 662 were isolated. Before fusion, mesophyll protoplasts of 662 were
irradiated with UV light at 254 nm and 0.07 J cm-2. Thus, the DNA of wild type 662 was
completely inactivated as it was shown for the irradiated protoplast control. The irradiated
protoplasts did not develop further in protoplast culture medium. Protoplast fusion was
performed using polyethylene glycol 6000. Already after one week, cell wall was re-
established and the cells proliferated well in protoplast culture medium. On solid media
protoplast calli developed shoots. Thirty-five plants were planted in soil. RAPD profiles of
twelve hybrid plants displayed characteristic DNA fragments of both protoplast donors. By
RFLP, the cytoplasmic genome of the regenerated plants was analysed. In six fusion plants
DNA recombination of mitochondrial and plastid genome occurred. By flow cytometry, the
DNA amount of the plants was investigated. Protoplast donor ‘Antik Pink’ and 35 fusion
plants had the same peak position indicating that only a small part of 662 DNA was
included in the asymmetric somatic hybrids. Plants flowered and 32 plants including nine
somatic hybrids formed seeds.
49
Species diversity and current status of Quercus L., representative genus
of natural flora of Crimea
In Crimean forests, these trees grow naturally: Q. pubescens, Q. petraea, Q. robur and
Q. petraea subsp. medwedievii. Oak formations occupy about 142 thousand hectares, or
54.7% of the total area of peninsula’s natural forests. The most extensive formation is the
stands of Q. petraea – 78.3% and Q. pubescens − 21.5% of the area of all oak trees. The
distribution of different types of Quercus and theirs satellites depends on climatic and soil
conditions. The stands of Q. petraea are not productive enough: this area is dominated by
plantations of IV-V bonitet classes. The age structure of natural forests of Q. petraea is
dominated by stands of VII-XI age classes, occupying 82.6% of the area of stands of this
species in Crimea. Age structure of forest stands of Q. pubescens, as well as Q. petraea is
characterized by maldistribution. The extremely limited number of plantings under 50
years testifies to the demographic imbalance of natural populations of Q. pubescens and low
level of natural revegetation processes. Currently, in the forest formations of Q. pubescens
in Crimea the low-tree stands of vegetative origin are dominating. In the existing situation,
plantings of Q. pubescens are losing their protective functions and degrading, especially in
the southern macroslope of the Main ridge of Crimean Mountains. Generally, the formation
of oak forests of Crimea is characterized by an unsatisfactory state, which is related to the
irrational forest management in the past. The issue that is important at the present time in
conservation of the natural plantations of Quercus genus in Crimea is a complex measures'
system building to improve the efficiency of their reproduction, using the biotechnology
methods.
Keywords: species diversity, oak, natural populations, productivity, age structure, seed,
reforestation.
50 50
Видовое разнообразие и текущее состояние представителей рода
Quercus L. природной флоры Крыма
51
The management of the growth of microplants of woody fruit crops
Sergey Kornatskiy
Peoples’ Friendship University of Russia, 6 Micklukho-Maklaya Str., Moscow, 117198,
Russian Federation; E-mail: vitrolab@rambler.ru
The success of the method of clonal micropropagation are obvious, but there is an
unresolved problem of effective growth of adapted microplants. Earlier we repeatedly
noted that microplants of such crops as sour cherry, sweet cherry, plum and garden
ashberry through 1-1,5 months after adaptation stop growing, fall into a state of rest, which
lasts for 3-7 months. The purpose of the research was to study the growth characteristics
of adapted microplants in simulating the seasonal rhythm of development under artificial
conditions. We suggested the reception of preliminary preparation of adapted micro-plants
for transplantation for growth. Microplants in a state of rest, were treated with low positive
temperatures + 2 ... + 6° C for 2.5 months in artificial conditions. Such influence allowed to
provide plants with active and effective development after creation optimal conditions for
vegetation. After 2-3 days, the plants had a synchronous onset of development, which
allowed a growth of 15-30 cm in length for 2-3 weeks in the initial plant height of 5-7 cm.
After this, a cyclic scheme was developed for alternating vegetation and rest periods in the
control of environmental factors. During the calendar year, we managed to realize growth
cycle 3 (1.5 months of vegetation + 2.5 months of rest) and almost completely reduce the
loss of material. The use of this scheme allowed to increase the microplants of the studied
crops to a height of 60-110 cm in the greenhouse in one year, which ensured their survival
in the field.
52 52
Управление ростом адаптированных микрорастений древесных
плодовых культур
С.А. Корнацкий
Российский университет дружбы народов, ул. Миклухо-Маклая, 6, г. Москва, 117198,
Российская Федерация; e-mail: vitrolab@rambler.ru
53
Genetic diversity of Tulipa suaveolens in the Crimea based on ISSR data
The research is partially funded by Russian Foundation of Basic Research under grant № 16-
04-00142.
54 54
Генетическое разнообразие Tulipa suaveolens в Крыму по данным
ISSR маркирования
55
Genetic diversity and population structure of the rare species, Artemisia
hololeuca detected by AFLP analysis
56 56
Генетическое разнообразие и популяционная структура редкого
вида Artemisia hololeuca на основании данных AFLP-анализа
57
Experimental variability of grapes in Vitaceae family based on the use of
allotetraploidy and in vitro embryo culture
Formation of grapes samples occurred in the course of natural evolution and is possible in
the process of experimental evolution under artificial generative hybridization. Crossing
forms of grapes with a different number of chromosomes suggest the need to combine
several methods that take into account the physiological characteristics of the initial forms.
An attempt was made to achieve intergeneric hybrids Vitis Ampelopsis and
Vitis Parthenocissus within the Vitaceae family using tetraploid genotypes of the initial
forms. In order to obtain tetraploid seedlings (diploid gametes), buds broken from
dormancy and flower clusters were treated with 0.5%, 1% and 2% colchicine for 5-14 days
before meiosis. The intergeneric incompatibility was circumvented by growing seedlings
from in vitro cultured embryos isolated from immature seeds for 40 days after pollination.
An intermediate dissection pattern of the leaf typical of the species of the genera Vitis and
Ampelopsis used in intergeneric hybridization by the allotetraploidy method was registered
in a number of hybrid seedlings. More than that, this is a proof of the biologically
determined possibility that forms from distant crosses may have emerged in nature in the
course of natural evolution. Due to the fact that it was more successful to obtain plants
from the hybridization of Vitis vinifera cultivars of the genus Vitis with the forms of
Ampelopsis genus species, in comparison with the crossing of the Vitis vinifera cultivars
with the forms of the Parthenocissus genus species, can be assume a closer genetic
relationship in the Vitaceae family between the genera Vitis and Ampelopsis, which are
closer in terms of the evolution of the grapes culture as a whole.
58 58
Экспериментальная изменчивость винограда семейства Vitaceae на
основе использования аллотетраплоидии и культуры эмбриоидов
in vitro
С.В. Левченко, В.А. Волынкин, В.А. Зленко, А.А. Полулях, В.В. Лиховской,
И.А. Васылык
Всероссийский национальный научно-исследовательский институт виноградарства
и виноделия "Магарач" ул. Кирова, 31, г. Ялта, 298600, Российская Федерация;
e-mail: svelevchenko@rambler.ru
59
Regeneration peculiarities of in vitro berry cultures
Russian Federation;
3FSFIS The Main Botanical Garden named after N. V. Tsitsin RAS, Botanicheskaya Str., 4,
In vitro collection of berry crops in Volgograd Regional Botanical Garden and Main
Botanical Garden named after N.V. Tsitsin of RAS has more than 300 accessions and consist
of representatives of the following families and genera: Grossulariaceae DC. (Grossularia
Mill., Ribes L.) Caprifoliaceae Juss. (Viburnum L., Lonicera L.), Rosaceae Аdans. (Rubus L.,
Cerasus Mill., Armeniaca Mill., Sorbus L., Prunus L.), Actinidiaceae Van-Tieghem (Actinidia
Lindl.). The optimal period of isolation of berry crop explants is the phase of the beginning
of active growth (April-May), while the yield of viable explants averaged 83%. To obtain a
sterile culture of fruit plants, in particular Actinidia, raspberries, honeysuckle, Lysoformin
3000 was used at a concentration of 5%, the exposure time was 3 min. When used, the
yield of sterile explants was maximum and was about 90%. The stage of micropropagation
used the following growth regulators combinations: 6-benzylaminopurine (6-BAP), 6-
benzylaminopurine ribozid, zeatin, kinetin – 6-furfurylaminopurine (KIN), kinetin ribozid,
2-isopentenyladenine (2-iP), thidiazuron (TDZ), Indole-acetic acid (IAA), indolebutyric acid
(IBA) and gibberellic acid (GA3). A comparative analysis of the study of the influence of
different cytokinins on representatives of the genus Actinidia showed that when using 2-iP
multiplication factor, A. kolomikta and A. arguta significantly exceeded the value of this
indicator in environments with 6-BAP. For A. polygama, the maximum value of the
multiplication factor on media containing 6-BAP (0.5 mg L-1) or zeatin (5.0 mg L-1) was
noted. At the same time, the content of zeatin in the medium increased the multiplication
factor in all studied Actinidia samples, simultaneously stimulating the formation of callus,
which subsequently slightly hampered the adaptation of regenerated plants in vivo. For
representatives of the genus Lonicera is quite reasonable use of cytokinins in combination
with auxins. The highest reproduction rate (18±0.2) was observed at 6-BAP 1.0 mg L-1 and
IBA 0.5 mg L-1. At the stage of rooting for Actinidia, optimal concentration is IAA 1.0 mg L-1;
for primocane raspberry – IAA 5.0 mg L-1; for honeysuckle – IBA 1.0 mg L-1. At the stage of
adaptation of berry crops, the use of two-stage adaptation is very effective. The yield of
adapted plants averaged 70-95%.
60 60
Особенности регенерации in vitro ягодных культур
61
Biological diversity of aromatic and medicinal plants in the Nikita
Botanical Gardens
This study was funded by the Russian Science Foundation, grant No. 14-50-00079
62 62
Биологическое разнообразие ароматических и лекарственных
растений в Никитском ботаническом саду
63
Genetic modification of jasmonate biosynthesis pathway in wheat alters
plant tolerance to necrotrophic fungi
Chemistry of the RAS, Prospect Nauki, 6, Puschino, Moscow region, 142290, Russian
Federation
The work was supported by the Russian Science Foundation, grant No 16-14-10155.
64 64
Изменение пути биосинтеза жасмонатов путем генетической
модификации пшеницы приводит к изменению устойчивости
растений к некротрофным грибам
65
Structure of the leaf blades in some horticultural plants cultured in vitro
Methods of plant tissue culture in vitro are widely used in modern fundamental and applied
researches in plant physiology, genetics, embryology and developmental biology. One of
the main topics is the investigations of plant assimilating organ structure, which is
associated with their successful adaptation in vivo. Thus, histological analyzes of the leaf
blades in a number of valuable essential oil, fruit and ornamental plants: Diospyros kaki
Thunb., Ficus carica L., Lavandula angustifolia Mill., Lavandula intermedia Emeric ex
Loiseleur, Rosa damascene Mill., Rosa chinensis, Chrysanthemum morifolium Ramat.,
Clematis L. и Canna hybridа hort. ex Backer. were carried out. The explants were cultured
for 12 months on the modified MS media in a growth chamber at 24±1°C, 16-hour
photoperiod and light intensity 37.5 μmol m-2s-1.
The leaf thickness in the in vitro propagated plants varied from 91 μm (R.
damascene) to 235 μm (C. hybridа). There was thin cuticle on epidermis surface (3-7 μm)
in Rosa damascene, Rosa chinensis, Lavandula angustifolia, Lavandula hybrida. and Ficus
carica. Multiple simple and glandular trichomes (81-303 μm) were noted on a leaf blade
surfaces in L. angustifolia, L. hybrida, F. carica, Chrysanthemum morifolium and Diospyros
kaki. Amphistomatic leaves were in Canna hybrida and Ch. morifolium, in the other
studied species, they were hypostomatic. The number of stomatal apparatus per 1 mm2
varied from 15 (C. hybridа) to 265 ones (R. damascene). Mechanical and vascular tissues
were well developed in all the studied cultivars. Differentiated palisade and spongy tissues
were characteristics of F. carica, R. damascene, R. chinensis, L. anustustifolia, L. hybrida,
some D. kaki and Clematis cultivars. Palisade index varied from 0.32 to 0.58. Homogeneous
mesophylls with large intercellular spaces was noted in C. × hybrida, most cultivars of
D. kaki, Ch. morifolium and Clematis. Thus, it has been identified a number of cultures
which under in vitro propagation demonstrated structural features typical for ex situ
plants, providing their better adaptation ex vitro and in vitro.
This study was funded by research grant No. 14-50-00079 of the Russian Science Foundation.
66 66
Структура листовых пластинок некоторых садовых растений,
культивируемых in vitro
И.В. Митрофанова, В.А. Браилко, С.В. Челомбит, Н.Н. Иванова, О.В. Митрофанова
ФГБУН «Ордена Трудового Красного Знамени Никитский ботанический сад –
Национальный научный центр РАН», Никитский спуск 52, пгт Никита, г. Ялта,
298648, Российская Федерация; e-mail: valentina.brailko@yandex.ru
67
Light intensity as the determining factor for in vitro regeneration in
some relic endemic species of the Crimea flora
One of the least studied aspects of the Crimean flora relict endemics propagation and
preservation in vitro is the effect of physical factors (light intensity and spectrum,
photoperiod) on the regeneration capacity. In the presented studies, the shoot nodal
segments of the species Lamium glaberrimum (K.Koch) Taliev, Scrophularia exilis Popl.,
Heracleum ligusticifolium M. Bieb., Lagoseris callicephala Juz., Lagoseris purpurea L. and
Silene jailensis N.I. Rubtzov were used. In the experiments, influence of white or red light
with the intensity 2,000; 3,000 and 4,000 lux with 16-h photoperiod and white light (2,500
lux) with 14-h photoperiod at the constant temperature (21-22°C) on regeneration
capacity were investigated.
Positive effect of the white light on the microshoot regeneration in Lam. glaberrimum,
Scr. exilis, L. callicephala, L. purpurea and S. jailensis was revealed at the light intensity of
2,000 lux and 16-hour photoperiod. However, in Lam. glaberrimum microshoot length, the
number of leaves and internodes were significantly higher in comparison with other species.
Under the same physical factors in the species Scr. exilis, the number of microshoots per
explant was significantly higher (18 ones). An increase in the white light intensity up to
3,000-4,000 lux caused leaf chlorosis in Lam. glaberrimum and Scr. exilis, as well as
microshoot lengthening, chlorotic leaves appearance and hydration in S. jailensis. Light
intensity 3,000-4,000 lux induced an increase in the number of adventitious shoots and
hydrated leaves in L. callicephala and L. purpurea. The red light with an intensity of 2,000-
4,000 lux caused leaf and microshoot hydration in Lam. glaberrimum and Scr. exilis. At light
intensity of 2,000 lux, the shoots of S. jailensis were thinned, with long internodes, at 3,000
and 4,000 lux intensity and they elongated greatly (had long internodes and chlorotic
leaves). Active formation of adventitious shoots under the red light and 2,000 lux intensity
was noted in L. callicephala and L. purpurea. Leaf yellowing and hydration were observed at
3,000-4,000 lux. Data obtained for H. ligusticifolium species are of great interest.
Regeneration frequency increased 2-2.5 times under the red light, 2,000 lux intensity and 16-
h photoperiod was demonstrated. An increase of the light intensity to 3,000-4,000 lux
resulted in the hydrated shoot and leaf formation.
Thus, original data on the development biology in six relict endemic species of the
Mountain Crimea flora have been obtained for the first time and it was demonstrated that the
physical factors of culture had a significant effect on the morphogenetic capacity realization
in vitro.
Keywords: rare species, regeneration capacity, light intensity, light spectrum, photoperiod, in
vitro.
68 68
Освещенность как определяющий фактор регенерации in vitro
некоторых видов реликтовых эндемиков флоры Крыма
О.В. Митрофанова, И.В. Митрофанова, Н.Н. Иванова, В.А. Браилко, Т.Н. Кузьмина,
Н.П. Лесникова-Седошенко, И.В. Жданова
ФГБУН «Ордена Трудового Красного Знамени Никитский ботанический сад –
Национальный научный центр РАН», Никитский спуск 52, пгт Никита, г. Ялта,
298648, Российская Федерация; e-mail: invitro_plant@mail.ru
69
Conservation and clonal micropropagation of rare endemic species
Aristolochia manshuriensis Kom.
In order to conserve plant biodiversity in vitro, methods have been successfully used in
recent years. A collection of rare plant species preserved in the form of meristem has been
created in Laboratory of Plant Biotechnology of the FSFIS The Main Botanical Garden
named after N. V. Tsitsin RAS. Aristolochia manshuriensis Kom. is a relict liana, endemic of
the Manchurian floristic region with limited habitat, gradually shrinking due to
anthropogenic impact. This species is listed in the Red Book of Russian Federation as
endangered one. A. manshuriensis hardly multiplies generative and vegetative ways, so to
obtain a sufficient amount of material for the further reintroduction purpose, clonal
micropropagation is used. For the representatives of the family Aristolochiaceae
(Aristolochia manshuriensis) at the first stage, the influence of the explant age and its
morphological features on the regeneration ability were investigated. As a result, it was
found that the explants taken from immature and virgin (not older than 4-6 years) plants
A. manshuriensis, characterized by a higher ability to proliferate shoots compared with
plants that have reached 12 years. The higher morphogenetic capacity was characterized
by the explants taken from 2-3-year-old plants. The highest multiplication factor
(14.84±0.8) was achieved on Murashige and Skoog medium with addition of 0.8 mg L-1 6-
BAP and 0.05 mg L-1 IAA. The optimum culture medium for rooting: ½MS containing 3.0 mg
L-1 IBA. The long-term storage in vitro not only allows to preserve of rare plant species, but
it is also a basis for studying the processes of morphogenesis and regeneration in tissue
culture including adaptation of microclones ex vitro.
70 70
Сохранение и клональное микроразмножение редкого эндемичного
вида Aristolochia manshuriensis Kom.
71
The increase of the effectiveness of small-fruit crop clonal
micropropagation by biophotonics
72 72
Повышение эффективности клонального микроразмножения
ягодных культур средствами биофотоники
73
Some biochemical features of Olea europaea L. frost resistance
74 74
Биохимические аспекты морозостойкости Olea europaea L.
75
The dynamics of the optimums of net-photosynthesis changes and
transpiration of Prunus laurocerasus L. and Viburnum tinus L. during a
vegetative period in the conditions of the Southern Coast of the Crimea
76 76
Динамика изменения оптимумов нетто-фотосинтеза, транспирации
Prunus laurocerasus L. и Viburnum tinus L. в течение вегетации в
условиях Южного берега Крыма
77
Strategy for sustentation of phytodiversity in the Crimea
The pace of phytodiversity reducing is impressive and the efforts to reign it in have not
been crowned with success yet. Some dystopian naturalists entertain an idea on waste of
money and irresistibility of attributable to evolution elimination of species, which have lost
their ecological niche. In opposition to them, realists try to take meaningful steps on
conservation for the species, which are threatened with extinction. To form a complete
meaningful program for conservation (strategy for sustentation) that could protect and
even recreate endangered species, it is critical to understand their populations’ condition
in wildlife, as well as to elicit how the populations react on different environmental
conditions and among other things on the climate transformation. It is important to know a
natural history and ecology of the species, as well. The strategic plan includes six
successive operations: 1. Estimation of a factor gradient length for the critical species
according to “Ecodata” database (297 vascular plants’ species are legislate under
guardianship of the law in the Crimea). “Ecodata” database contains the unified-standard
information on the state of plants’ species on the environmental factors’ gradients:
luminance-shadowing, a thermal regime, an ombrogenious regime, a cryogenic regime,
continentality, a moisture variability, a substrate acidity, an anionic structure,
calcareousness, a nitrogen content, a substrate granulometric texture; 2. Description of
critical species communities in wildlife (in situ); 3. Estimation species packaging density on
the environmental factors’ gradients with the use of an original program «Pover», which
allows to estimate wildlife habitual areas’ capacity and spot an optimum point position.
The species packing factor on short gradients is in evidence higher than that on the long
ones. A vector dimension is a length of a realized gradient (an amount of occupied with
gradations) of the studied factors-conditions and factors-sources shows an existential
source in the ecotope; 4. Elimination of a biological material for domestication (ex situ);
5. Propagation in the cultivar with subsequent adaptation to environmental conditions;
6. Repatriation to preselected ecotopes with a convenient ecological niche.
This study was funded by a research grant № 14-50-00079 of the Russian Science Foundation.
78 78
Стратегия поддержания фиторазнообразия в Крыму
79
Reproductive peculiarities of some hybrid tea-roses in the collection of
the Nikita Botanical Gardens
There are about 250 cultivars of the most popular in horticulture garden group of hybrid
tea-roses in the collection of the Nikita Botanical Gardens (NBG). The study of the
reproductive peculiarities of hybrid tea-roses allows supplementing their introductive
estimation in the conditions of the Southern Coast of the Crimea because the ability to form
full-blown fruits and seeds is one of the introduction success criteria. What is more the
knowledge on the ability of cultivars to form fruits and seeds as well as their pollen quality
is needed to involve the cultivars into hybridization as father- or mother seed parent forms.
The comparative study of the pollen quality of 100 highly decorative hybrid tea-roses by
the method of pollen grains staining with acetocarmine has been done. The pollen has been
gathered during the first blossoming phenological phase. It has been elucidated that the
studied cultivars are essentially different in the quantity of normally formed (being
stained) pollen, which is from 5 to 81.9%. As the results, the collection cultivars have been
enrolled by the quantity of morphologically full-blown painted pollen into five groups: up
to 20% (16 cultivars), from 20.1 to 40% (22 cultivars), from 40.1 to 60% (43 cultivars),
from 60.1 to 80% (15 cultivars), and more than 80.1 % (two cultivars). 60 cultivars, which
belong to three last-named groups, are recommended to use as father seed parent forms
while hybridization. Having studied fructification of hybrid tea-roses cultivars, it has been
determined that 32 cultivars do not set fruits, 24 cultivars set singular fruits and only 44
set fruits massively and are recommended to use as mother seed parent forms while
hybridization.
This study was funded by a research grant No. 14-50-00079 of the Russian Science
Foundation.
Keywords: pollen, fructification, hybridization, father seed parent form, mother seed
parent form
80 80
Репродуктивные особенности некоторых сортов чайно-гибридных
роз коллекции Никитского ботанического сада
81
Essential oil characteristics and expression analysis of scent-related
genes in four landraces of Damask rose
The main volatile compounds emitted from flower of Rosa damascena are monoterpenes.
To evaluate the essential oil content, flower physiochemical characteristics and gene
expression, this experiment was conducted on four landraces of Damask rose. The cDNA of
GGPPS gene was isolated from this plant applying nested-PCR method. This gene with
accession No. KX661005, encodes for a putative protein of 365 amino acid. Quantitative
real-time PCR analysis of GGPPS, DXR and CCD1 was done on mRNA samples extracted
from 3 floral developmental stages (A, B and C) of four landraces of damask roses including
‘Ghalhar’, ‘Lavasanat’, ‘Kamoo’ and ‘Minab’. Results indicated difference in expression levels
of genes and floeral stages, as the lowest level expression was revealed for GGPPS gene,
compared with those of genes CCD1 and DXR. In addition to molecular study, physiological
and phytochemical parameters were characterized focusing on relationship between main
volatile compounds and colors. Varying petal color from light pink to deep pink, the light
color flowers showed higher carotenoids and lower anthocyanin contents. During flower
development from A to C stages, main monoterpenes compounds likes citronellol, geraniol
and neral were increased till stage B, then reduced at stage C. However, these results
suggested that GGPPS as a key gene could be involved in monoterpenes biosynthesis and
could be important to enhance main volatile production.
82 82
In vitro shoot multiplication of an ornamental aquatic plant, Lobelia
cardinalis L.
The shoots of an ornamental Aquatic Plant, Lobelia cardinalis L. from ex vitro were surface
sterilized using 4% Clorox® (5.25% sodium hypochlorite, NaOCl) for 10 min followed by
rinsing three times with sterile distilled water. They were again surface sterilized for
another 2% Clorox® for 5 min. All shoot tip explants were cultured on MS (Murashige and
Skoog, 1962) medium for multiple shoot regeneration. The media were further augmented
with various concentrations of growth regulators namely N6-benzylaminopurine (BAP; 0.0,
1.0, 2.0 or 3.0 mg L-1) and α-naphthaleneacetic acid (NAA; 0.0, 1.0, 2.0 or 3.0 mg L-1).
Presence of BAP in the medium significantly increased shoot numbers. MS medium
supplemented with 1.0 mg L-1 BAP was significantly different compared to other
treatments, with the highest number of 5.8±0.74 shoots per explant and shoot formation at
100% after 60 days of culture. The result revealed that MS medium supplemented without
plant growth regulators was significantly different compared to other treatments, with the
highest number of roots per shoot 11.4±2.00 roots per explant and root length at
22.37±1.67 mm. Rooted plantlets were subsequently hardened, acclimatized and
successfully established in field with 95% survival rate. Acclimatized plantlets of Lobelia
cardinalis L. were vigorously growing without any symptoms of diseases even after 30 day
of acclimatization.
83
Macro and trace element accumulation by species of genus Artemisia at
the Southern Coast of Crimea
Gardens – National Scientific Center of the RAS", Yalta, 298648, Russian Federation;
E-mail: oksana_shevchuk1970@mail.ru;
2Federal Scientific Center of Vegetable production, VNIISSOK, Selectsionnaya Str., 14,
84 84
Накопление макро- и микроэлементов видами рода Artemisia в
условиях Южного берега Крыма
О.М. Шевчук1, Н.А. Голубкина2, Л.А. Логвиненко1, А.В. Молчанова2, Ю.В. Плугатарь1
1ФГБУН «Ордена Трудового Красного Знамени Никитский ботанический сад –
Национальный научный центр РАН», Никитский спуск 52, пгт Никита, г. Ялта,
298648, Российская Федерация; e-mail: oksana_shevchuk1970@mail.ru
2ФГБНУ Федеральный научный центр овощеводства, пос. ВНИИССОК,
85
NBS-LRR resistance genes polymorphism in apple (Malus domestica
Borkh.) landraces inferred from NBS-profiling
Anna Shlyavas1, Aya Trifonova2, Liliya Dedova2, Ksenia Boris2, Alexander Kudryavtsev2
1FRC The N. I. Vavilov All-Russian Institute of Plant Genetic Resources (VIR), Bol’shaya
86 86
Полиморфизм NBS-LRR генов устойчивости сортов яблони (Malus
domestica Borkh.) народной селекции по данным NBS-профайлинга
А.В. Шлявас1, А.А. Трифонова2, Л.В. Дедова2, К.В. Борис2, А.М. Кудрявцев2
1Всероссийский институт генетических ресурсов растений имени Н.И. Вавилова,
87
The role of introduction in increasing peach (Persica vulgaris Mill.)
genetic diversity at the Nikita Botanical Gardens collection
By means of introduction and breeding, the Nikita Botanical Gardens created a gene pool of
peach, consisting of 559 cultivars, which comprises 288 cultivars by NBG (51.5%), 101
cultivars from USA (18.1%), 21 from China (3.8%), 55 (9.8%) from Central Asia and
Transcaucasia, eight from Canada (1.4%), 57 (10.1%) from 14 European countries and 29
(5.2%) cultivars of unknown origin. Ecogeographical groups and ecotypes were classified
in accordance with fruit quality (consistency and coloration of flesh, coating coloration,
weight, stone freeness, ripening time and taste). There was detected the incidence of each
trait in ecogeographycal groups and ecotypes. North Chinese and Iranian ecogeographical
groups comprised mostly cultivars of European (168 and 182, respectively) and American
(32 and 77) ecotypes. The research of 128 mid-season cultivars (40.8%) showed that most
of them belonged to European ecotype of North Chinese (75) and Iranian (74)
ecogeographical groups. 38.5% of cultivars of the peach collection of the NBG present a
good separable stone. 126 of them belonged to Iranian ecogeographical group and 89 to
North Chinese one. The largest fruit cultivars belonged to European ecotype of North
Chinese (31) and Iranian (46) ecogeographical groups. In Iranian ecogeographical group,
the part of cultivars with harmonious taste of fruits accounted for about 60%, in the North
Chinese one – 40%. These cultivars belonged mostly to European and American ecotypes.
Within the framework of breeding programs, 46 cultivars have been developed and
included into commercial range of peaches by the use of 22 initial cultivars. So far, the
Nikita Botanical Gardens obtained the RF patents for 30 cultivars of peach.
This study was funded by a research grant No. 14-50-00079 of the Russian Science
Foundation.
88 88
Значение интродукции в расширении генетического разнообразия
генофонда персика (Persica vulgaris Mill.) коллекции Никитского
ботанического сада
89
Influence of ecological and geographical conditions on the biochemical
potential of the genotype of golden currant
Golden currant is a valuable food culture, which has a wide ecological plasticity, which
allows to cultivate it in the conditions where other types of currants feel oppressed. Golden
currant berries contain sugar (6.3-17.0 %), dry matter (17-25%), vitamin C (23.2-125
mg/100 g), pectin (0.6-2.9%), organic acids (up to 2.1%), anthocyanins (200-470 mg/100
g). According to the content of carotene in berries (8-19 mg/100 g in terms of β-carotene),
golden currant takes the first place in the genus currants, the maximum content of these
biologically active substances is typical for the most dark-colored forms.
It was made a long-term (2010-2016) comparison of three golden current cultivars:
‘Venus’, ‘Lyaysan’ and ‘Shafak’ in the Belgorod region (a temperate continental climate) and
the Republic of Bashkortostan (a continental climate) in Russia. The change in temperature
and water conditions lead to changes not only in the content of fruit’s biological substances,
but also in size. With an increase in humidity by 16% and decrease in temperature by 13°C
(Bashkortostan), the content of dry soluble substances increased from 1.3% (‘Lyaysan’) to
11.9% (‘Venus’) and vitamin C increased in the cultivars ‘Venus’ (4.5 mg/100 g) and
‘Shafak’ (11.8 mg/100 g). For the cultivar ‘Lyaysan’, the content of vitamin C decreased by
9.1 mg/100 g, also average sugar content among the cultivars decreased by 2%. The
influence of ecological and geographical conditions on the content of anthocyanins and
carotenoids in the studied fruits is not significant.
90 90
Влияние эколого-географических условий на биохимический
потенциал генотипа смородины золотистой
91
SSR-fingerprinting and genetic relationship study of modern peach
cultivars from the germplasm collection of the Nikita Botanical Gardens
Ivan Suprun1, Anatoliy Smykov2, Iliya Stepanov1, Sergey Tokmakov1, Olga Fedorova2, Iuliia
Ivashchenko2
1Federal State Budget Scientific Institution “North Caucasian Federal Scientific Center of
Gardens – National Scientific Center of the RAS", Yalta, 298648, Russian Federation
A sub collection of 40 peach (Prunus persica (L.) Batsch) cultivars from the collection of
genetic resources of the Nikita Botanical Gardens was evaluated by microsatellite DNA-
markers for estimation of genetic diversity and relationship within studied group of
cultivars. Totally 12 SSR markers were used for genotyping. Markers showed different level
of polymorphism: from three (for markers UDP98-410 and BPPCT028) to nine
(BPPCT017) alleles per locus with mean value 5.417 alleles per locus while number of
effective alleles varied from 3.970 to 1.261 with mean value 2.512. Expected and observed
heterozygosity varied within the ranges 0.207–0.748 and 0.075–0.875, respectively.
According to the level of polymorphism, several markers are recommended as perspective
for using in the further studies of genetic diversity. For all the studied cultivars unique SSR-
fingerprints were obtained. According to results of UPGMA–analysis, three main groups of
cultivars may be determined. By the way, few cultivars were separated into out groups.
These are: Chinese cultivar ‘Dzyu-yus-tszyuy’, which was determined as the most
genetically distanced from all of the cultivars and additionally two cultivars – ‘Dostoyniy’
(breeding of the Nikita Botanical Gardens) and ‘Andrey Lupan’ (Moldavian breeding) which
have specific fruit traits – highly crispy and low melting flesh. Distribution in the main
three clusters in some cases is corresponding with genealogy of cultivars. The obtained
data has showed genetic relationship among studied peach cultivars, that represent the
modern assortment from the collection of the Nikita Botanical Gardens.
92 92
SSR-фингерпринтинг и изучение генетических взаимосвязей
современных сортов персика из коллекции генофонда Никитского
ботанического сада
И.И. Супрун1, А.В. Смыков2, И.В. Степанов1, С.В. Токмаков1, О.С. Федорова2,
Ю.А. Иващенко2
1Федеральное государственное бюджетное научное учреждение «Северо-Кавказский
93
Micropropagation of Larix sibirica L. by somatic embryogenesis in
culture in vitro
Branch of the RAS - Division of Federal Research Center «Krasnoyarsk Scientific Center of
the Siberian Branch of the RAS», 50/28Akademgorodok, Krasnoyarsk, 660036, Russian
Federation
2 Laboratory of Plant Physiology, Institute of Biology of Ufa Federal Research Centre of
The isolated megagamethophytes and zygotic embryos at the stages of cotyledon initiation
were used as the explants for induction of somatic embryogenesis of Siberian larch (Larix
sibirica L.). The explants were cultivated on AI medium (patent №2456344
http://www.freepatent.ru/patents/2456344). The 42 proliferating embryogenic cell lines
(Cls) of Siberian larch have been received in different years (2008-2017) from two
genotypes No. A4 and No.10 as a result of opened and controlled pollination. Produced Cls
of L sibirica. were characterized by high embryogenic productivity. But CLs were initially
different in embryo production. Some CLs produced more somatic embryos than others,
under the same culture conditions. In highly productive (Cl4) embryos maturated and
germinated more actively. CL5 was a special case. Although somatic embryos did occur in
it, they did not maturate on the medium with ABA. According to immunohistochemical
staining of the cells, the globules contained IAA, zeatin and ABA, which varied among
different Сls and absent in embryonic tube of suspensor. Microsatellite analysis of
proliferating cell lines of L. sibiricah have shown weak allelic variability. Multiplication of
somatic embryos actively passed as a result of cleavage of embryos, suspensor budding and
embryonal tubes of suspensor proliferation. Mature somatic embryos outcome from the
primary number of globular somatic embryos in different CLs ranged from 0.6% to 33%.
Germination of somatic embryos was carried out on a non-hormonal ½AI medium, The
most regenerants are characterized by the formation of callus in the area of the hypocotyl
and between hypocotyl and root. But they are regenerants (Cl4) with good roots. For
adaptation, regenerants were planted in sterile soil in a growth chamber, then in a
greenhouse and further into the forest nursery soil, where cloned seedlings actively grow
for seven years. Genotyping of microsatellite loci in seedlings showed full compliance with
their CL6 from which they were derived.
The work is executed at financial support of RFBR grant No. 15-04-01427 and grant No.18-
54-00010 Bel_a.
94 94
Micropropagation of berry crops for creation of germplasm cryobanks
Timur Turdiev, Zinat Mukhitdinova, Sergey Frolov, Akzhol Nogaibayev, Irina Kovalchuk
Institute of Plant Biology and Biotechnology, 45 Timiryazev Str., Almaty, 050040,
Kazakhstan; E-mail: turdievtt@mail.ru
This study was funded by Ministry of Education and Science of the Republic of Kazakhstan, project
No. AP05131764.
95
Magnetic-pulse processing at micropropagation and cleaning up of fruit
and small fruit crops from viruses
Mikhail Upadyshev, Ivan Kulikov, Vladislav Donetskich, Anna Petrova, Klavdia Metlitskaya
and Galina Upadysheva
All-Russian Horticultural Institute for Breeding, Agrotechnology and Nursery,
Zagorievskaya Str., 4, Moscow, 115598, Russian Federation; E-mail: upad8@mail.ru
Outer low-frequency magnetic fields can have influence on the process of plant growth,
development and cleaning up from viruses. The effects depend on the magnetic field
characteristics, on the biological peculiarities of the culture and the kind of pathogen. The
purpose of the research was to study the magnetic-pulse processing action on the
micropropagation and cleaning up process from harmful viruses of apple, pear, sweet
cherry and raspberry plants. The explants cultivated on Murashige and Skoog culture
medium, and infected by the viruses have been processed with the help of the
programmable devices designed according to our own construction (patents of the Russian
Federation No.2573349 and No.2652818) by the magnetic induction impulses with the
frequency from 0.5 to 130 Hz. The microplants testing was fulfilled by the ELISA-test. The
usage of magnetic processing on apple and pear plants provided 86% of the explant
cleaning up from the complex of viruses ASPV, ASGV, ACLSV and ApMV, on sweet cherry
plants – up to 29% from the complex of viruses PPV, PNRSV and PDV. On raspberry plants
the magnetic processing caused virus RBDV inhibition, on raspberry-blackberry hybrid –
from viruses RBDV and TBRV (the output of virus-free plants – up to 67%). The magnetic
processing encouraged the improvement of explant development at the micropropagation
stage: the microshoots length grow up in 1.5-2.9 times, their number – in 1.3-2.7 times
depending on the crops. At the stage of rooting ability, the magnetic processing increased
this ability for the microshoots in 2-3.3 times, the roots number and length – in 1.6-3.1
times, the microplant acclimatization in non-sterile conditions – on 7-27%. The advantages
of the magnetotherapy are the absence of phytotoxicity, universality, higher output of
health plants, processing automatisation and ecological safety.
96 96
Магнитно-импульсная обработка при микроразмножении и
оздоровлении от вирусов плодовых и ягодных культур
М.Т. Упадышев, И.М. Куликов, В.И. Донецких, А.Д. Петрова, К.В. Метлицкая,
Г.Ю. Упадышева
Федеральное государственное бюджетное научное учреждение «Всероссийский
селекционно-технологический институт садоводства и питомниководства»,
ул. Загорьевская, 4, г. Москва, 115598, Российская Федерация; e-mail: upad8@mail.ru
97
Biodiversity of distant hybrids of cereals in the collection of the Tsitsin
MBG RAS
Vladimir Upelniek, Andrey Fisenko, Lubov Ivanova, Lubov Gluhova, Nina Kuzmina,
Pavla Loshakova, Sergey Zavgorodny, Sergey Gradskov
FSFIS The Main Botanical Garden named after N. V. Tsitsin RAS, Botanicheskaya Str., 4,
Moscow, 127276, Russian Federation; E-mail: vla-upelniek@yandex.ru
In Main Botanical Garden named after N.V. Tsitsin RAS more than 70 year scientists have
been working on preservation, studying and involvement the cultural and wild-growing
flora representatives of the cereal family in breeding process. The creation of distant
hybrids and their studying allow to solve not only fundamental problems, but also to obtain
new forms and species for breeding practice and production. In that regard, distant
hybridization offers exciting possibilities for using genome potential of wild plant species.
The creation of new synthetic species Trititrigia cziczinii Tzvel. (2n=56) can be considered
as the obvious success. Trititrigia cziczinii Tzvel. (2n=56) is the new species, based on
crossbreeding of varieties of different wheat species with wheatgrass species. The genome
is presented by 42 wheat chromosomes and 14 wheatgrass chromosomes. This species
representatives are characterized by new attributes, non-typical for common wheat
(perennity and ability to grow shoots after mowing) and also by bread-making quality
(protein 21%). The modern collection includes more than 250 Trititrigia cziczinii Tzvel.
forms. The wheatgrass collection involves both initial natural populations of different
geographical origin and set of two species inbred lines (in total not more than 5,000 plants)
Elytrigia elongata (Host) Nevski, Elytrigia intermedia (Host) and of two reciprocal hybrids:
× Elytrigia intel Cicin. and Elytrigia elint Cicin. Apart wheatgrass, collections of other wild-
growing species and genera of the cereals family are preserved and studied: wild
ruttishness (Elymus dahuricus Turcz. Ex Griseb, El. canadensis L., El. pungens (Pers.)
Melderis., El. farctus (Viv.) Runemark. ex Melderis), fescue (Festuka rubra L., F. ovina L.),
bluegrass (Poa annua L., P. pretense L.), goat grass (Aegilops tauschii Coss., Ae. longissima
Schweinf. & Muschl., Ae. speltoides Tausch.) and lyme grass (Léymus arenárius L.). The
collection of wheat-wheatgrass hybrids totals 120 samples. The collection of spring and
winter tritici type wheat-wheatgrass hybrids includes 520 samples. The new forms of
distant hybrids obtained in recent years by the hybridization of triticale and trititrigia
(Triticosecale × Trititrigia) are of the especial interest. A large number of essential for
breeding samples makes this collection really unic not only from the point of view of
preserving the genetic diversity of cereals, but also as an important source of economic
characters and gene associations in breeding practices and genetic studies.
98 98
Биоразнообразие отдаленных гибридов злаков в коллекции ГБС
РАН
В.П. Упелниек, А.В. Фисенко, Л.П. Иванова, Н.П. Кузьмина, П.О. Лошакова,
С.В. Завгородний, С.М. Градсков
Федеральное государственное бюджетное учреждение науки Главный ботанический
сад им. Н.В. Цицина Российской академии наук, Ботаническая ул., 4, г. Москва,
127276, Российкая Федерация; e-mail: vla-upelniek@yandex.ru
99
Cytogenetics, phenology and development of biotechnological methods
for the conservation of Jatropha costaricensis
Marta Fermina Valdez1, Elmer García Díaz1, Andres Gatica-Arias1, Pablo Bolaños-Villegas2,
Nien Weng-Huang3
1School of Biology, University of Costa Rica, Ciudad Universitaria Rodrigo Facio, San Pedro
Costa Rica
3Faculty Pharmacy, University of Costa Rica, Ciudad Universitaria Rodrigo Facio, San Pedro
The genus Jatropha belongs to the Euphorbiaceae family and comprises 3 500 species. The
genus has a cosmopolitan distribution, including species native to Mesoamerica. Jatropha
plants are non-food oleaginous crops capable of adapting to different edaphic, ecological
and climatological conditions. Jatropha species found in Costa Rica include J. costaricensis,
J. curcas, J. integerrima, J. podagrica and J. stevensii. The species J. costaricensis was
described in 1978 and is an endangered species with only three populations, which are
found in the Province of Guanacaste, Costa Rica. It is not known to occur in other countries.
The aim of this project is to study the cytogenetics, phenology and phytochemical profile of
J. costaricensis, as well as to develop biotechnological methods for the establishment of cell
cultures in vitro, in order to allow massive micropropagation of plants, via organogenesis,
for their conservation. Also, it is of great importance to explore the potential of this wild
species in relation to an eventual capacity for the production of natural compounds for
pharmaceutical use.
The species was described as dioecious, but our preliminary observations indicate
that individuals having only male flowers coexist with trees having only female flowers, but
also with monoecious trees. The analysis of the lipid profile of J. costaricensis showed a
total of 12 fatty acids, with an overall lipid content of 38.5%, and an average seed weight of
1.425 g. These values are much higher than those of the commercial species J. curcas,
described in the literature: four fatty acids, 40% content and an average seed weight of 0.8
g. J. costaricensis has a high potential both as a genetic resource for the commercial variety
of the genus, J. curcas, and in relation to its own characteristics that make it very promising
from the pharmaceutical and cosmetically point of view.
100 100
Ontomorphogenesis of axillary meristem-derived micropropagated
plants of Rosa canina rootstock
Problem of garden roses reproduction in Western Siberia is connected with the absence of
seed plantations of winter-hardy rose-rootstocks. In CSBG SB RAS (Novosibirsk) a
comparative study of the plants ontomorphogenesis of the selected highly winter-hardy
Rosa canina form obtained by clonal micropropagation, seeds and green cuttings was
conducted in 2013-2018.
Primary explants for in vitro multiplication were meristems with two leaf primordia,
isolated from the axillary buds of annual shoots. The initiation of shoot formation was
carried out in MS liquid medium supplemented with 100 mg L-1 glutathione, followed by
cultivation on MS induction medium with 2.0 mg L-1 BA and 1.0 mg L-1 IAA. At the
micropropagation stage, the maximum number of microshoots per explant (7.5 ± 0.7) was
registrated on medium with 1 mg L-1 BA. For rooting of the microshoots, ½ MS hormone-
free medium supplemented with 1.0 mg L-1 IAA was used. The regenerants were
acclimatized in containers with sterile sand, followed by transferring into pots containing a
peat with perlite, mold, sand and coconut substrate. To date, in vivo conditions, the
ontogenesis of R. canina of microclonal origin, including three ontogenetic states of the
pregenerative period and two states of the generative period, has been described for the
first time. In the selective form studied, all the shoot formation systems (SFS) were
preserved completely. They formed fruit shoots with 1-5 gipantia with full fruit-nuts.
The technology of clonal micropropagation of selected forms of rootstock intended
for seed plantations is effective for accelerated reproduction, since there is no need in
stratification, and there is no vulnerable seedling in outdoor conditions. In addition, in vitro
technique used makes it possible to obtain a clonal stock material regardless of the season,
unlike green cuttings, which is important for conditions with a short vegetative period.
101
101
Онтоморфогенез подвоя Rosa canina L., полученного в результате
микроразмножения из меристем пазушных почек
102 102
Antioxidant activity of Solidago L. complex
The total antioxidant activity was evaluated by DPPH method (Brand-Williams, 1995) on
the dry plants (leaves and inflorescences). We compared 13 samples grown from seeds at
the experimental plots in the vicinity of Moscow: 1) native Solidago virgaurea L.; 2)
invasive S. altissima L. (=S. canadensis s.l) (Hungary); 3) invasive S. altissima (Belarus); 4)
invasive S. canadensis L. s.str (Hungary); 5) invasive S. gigantean Aiton (Hungary); 6,7)
hybridogenous × S. niederedery Khek. (=S. virgaurea × S. canadensis); 8-11) hybridogenous
× S. snarsksii Gudžinskas & Žalneravičius (=S. virgaurea × S. gigantea); 12) invasive
S. graminifolia (L.) Nutt. (=Euthamia graminifolia) (Belarus); 13) alien S. graminifolia (L.)
Nutt. (Brno). The antioxidant activity of alcohol extracts for all specimens was very high
and comparable to that of Juniperus sabina (Elisovetcaia et al., 2018) or Tussilago farfara
(Ivanišová et al., 2016). The total antioxidant activity in the leaves had 75.28-92.77%
(methanol extracts), 44.34-91.52% (ethanol extracts) and 21.55-68.51% (water extracts).
The total antioxidant activity in the inflorescences was higher: 90.77-94.07% (methanol
extracts), 90.20-95.66% (ethanol extracts) and 37.69-64.47% (water extracts). Alcohol
extracts in all experiments had 2.5-3.0 times higher antioxidant activity than aqueous
extracts. The lowest antioxidant activity in alcohol extracts was shown by S. snarsksii No.
11 (leaves), and in the aqueous extract – S. snarsksii No. 10 (leaves). This taxon, in general,
demonstrates the greatest variability of characters, which apparently is a reflection of its
hybridogenic nature. The alcohol extracts from the leaves of both samples of S. graminifolia
also show reliably lower antioxidant activity (84 vs. 91% in methanol and 85 vs. 90% in
ethanol), which to some extent is an indirect confirmation of the correctness of its transfer
to the genus Euthamia.
Consequently, alien Solidago species, as well as their hybridogenic taxa (excluding
S. snarsksii), are similar in antioxidant activity to the native S. virgaurea included in the
official pharmacopoeia. In this way, alien Solidago taxa have a potential source of useful
bioactive compounds.
103
103
In vitro conservation of essential oil rose cultivars
Gardens – National Scientific Center of the RAS", Yalta, 298648, Russian Federation;
2Federal State Funded Institution of Science “Research Institute of Agriculture of Crimea”,
The effect of explants сonservation in essential oil rose cultivars (Rosa spp.) during 1 year
on their morphometric and physiological parameters during preservation (under
conditions of slow growth) and after transfer to standard in vitro culture has been
investigated. The segments of the microshoots (5-7 mm) of cultivars ‘Raduga’, ‘Lany’,
‘Lada’, ‘Michurinka’, and ‘Festivalnaya’ were kept at 4-6°С and light intensity 1,000 lx.
During storage, viability of the explants of all investigated genotypes was at a high level
(90-100%). Herewith, depending on a cultivar, in 85.7-100% of explants, a slow
development (formation of new leaves, additional buds and shoots) was observed. The best
growth of adventitious shoots was in the cultivars ‘Festivalnaya’ and ‘Lany’ (1.8 and 2.1
pieces per explant, respectively). At low temperature, the work of photosystem-2 in the
leaves of all cultivars was reduced: the relative photosynthetic activity was 0.41-0.50 a.u.
At the same time in control (during а clonal micropropagation in meristem culture) in the
studied cultivars shoots the active light conversion was observed, which indicated normal
functioning of their assimilation apparatus. After one year of conservation, explants were
transferred to a fresh culture medium and cultured at 24-26°C with light intensity 2,000-
3,000 lux under 16-h photoperiod. For all the cultivars within a month, active shoot growth
and adventitious shoot formation (from 2.1 to 4.6 pieces per explant, respectively) were
observed. The multiplication index varied from 4.2 (‘Lada’) to 7.6 (‘Festivalnaya’). Some
morphometric parameters were on the control level, and for ‘Festivalnaya’ and 'Lany' even
higher. The analysis of the parameters of chlorophyll fluorescence induction of plant
material after transfer to standard in vitro culture conditions indicates restoration of
production processes. The cultivars ‘Raduga’ and ‘Michurinka’ showed a slight photo
inhibition.
This study was funded by a research grant No. 14-50-00079 of the Russian Science
Foundation.
104 104
Сохранение in vitro сортов эфиромасличной розы
Национальный научный центр РАН», Никитский спуск 52, пгт Никита, г. Ялта,
298648, Российская Федерация;
2ФГБУН «Научно-исследовательский институт сельского хозяйства Крыма»,
105
105
Integration of molecular and geographical data analysis of Iranian
Prunus scoparia populations in order to assess genetic diversity and
conservation planning
106 106
Technology of sugar beet dihaploid lines development
The methods used to create homozygous lines of sugar beet dihaploids when cultivating
unfertilized ovules under in vitro conditions are represented. At the first stage, use of
phenotype traits of seed-bearing plants, morphological and cytological features of formed
buds and unfertilized ovules is of vital importance to induce haploids. The main part of the
technology is devoted to presentation of the two-stage method for cultivation of isolated
ovules using liquid and agar-like consistencies of Gamborg’s culture medium (В5) which
regulates explants’ supply with hormones and nutrients. Identifying characteristic of this
method is a culture medium consistence change that allows differentiation of cultured
tissues in liquid phase and, when transferred to agar-like medium, induction of haploid
regenerants formation. Hormonal composition of Gamborg’s culture medium is an
important factor that effectively controls direction of morphogenetic development in
isolated ovules through direct regeneration (embryogenesis) or via callus
(gemmorhizogenesis) that testifies to totipotency of both sexual and somatic explant cells.
When producing lines of doubled haploids, stabilizing selection promotes detection of
valuable morphological characteristics in regenerants. Last stage is rooting of dihaploid
regenerants which success is determined by their transplanting to a new culture medium
with changed composition of auxins. Microclones with vigorous root system and well
developed leaf surface are used to obtain planting material. The method of induced
parthenogenetic plant selection includes cytophotometric ploidy evaluation from the
content of nuclear DNA at the stabilization stage of haploid regenerants and after
diploidization. Molecular-genetic studies of amplified DNA fragments of mitochondrial
genome allow genotyping of regenerants according to sterile and fertile cytoplasm types.
Final processes of the technology are: transfer of the rooted microclones with vigorous root
system and well-developed leaf surface to a greenhouse, growing seed-bearing plants from
them and obtaining seeds.
107
107
Технология создания дигаплоидных линий сахарной свеклы
108 108
Detection and partial molecular characterization of viruses
infecting chrysanthemum in Russia
1 Federal State Funded Institution of Science "The Labor Red Banner Order Nikita
Botanical Gardens – National Scientific Center of the RAS", Yalta, 298648, Russian
Federation;
2 Lomonosov Moscow State University, Моscow, 119234, Russian Federation;
E-mail: s-chirkov1@yandex.ru
This work was funded by the Russian Science Foundation, grant No 14-50-00079.
109
109
Обнаружение и частичная молекулярная характеризация
вирусов хризантемы в России
Национальный научный центр РАН», Никитский спуск 52, пгт Никита, г. Ялта,
298648, Российская Федерация;
2Московский государственный университет имени М.В. Ломоносова, г. Москва,
110 110
N-Acethylcysteinate- 2-ethyl-6-methyl-3-hydroxypyridine as a plant
growth regulator
111
111
N-ацетилцистеинат 2-этил-6_метил-3-гидроксипиридин в
качестве регулятора роста растений
112 112
The Nikita Botanical Gardens ornamental plants' collections biodiversity
extension
The collection of 8 ornamental plants in the Nikita Botanical Gardens (garden roses, irises,
daylilies, tulips, lilacs, cannas, clematis and chrysanthemum) account for 2,456 species,
cultivars and forms. The collections’ renewal is carried out by introduction, and also by
creating new domestic cultivars. A wide range of morphological and biological
characteristics qualitative variability within the species or the genus in each is represented
by the time being. The collections’ cultivars belong to different groups according to their
ornamental and biological characteristics: coloring, doubling, a flower form, flowering
duration, flowering term. The cultivars, which have been introduced over the last years, are
got through the process of introductive testing and are bred to put in the collections. As the
result of the introductive work, the collections’ biodiversity has been expanded constantly.
For the time being, the study of the new cultivars for the garden groups – hybrids
Hulthemia, spray roses, patio roses, the group of English roses’ cultivars by D. Austin’s
introduction, Atragene, Flammula, Tangutica, Texensis clematis’groups and Veridiflora
tulips’ garden class is being done. The cultivars with the new characteristics are prepared
to put in the collection: polypetalous lilac cultivars, garden roses’ cultivars with a coffee
and blue coloring, with carved edges of petals, daylilies with a spider flower form, hybrid
irises with a spatial continuation of a beard that looks like a horn and a petaloid,
chrysanthemum cultivar with a filiform spider narrow-tubular inflorescence form, garden
cannas with a dark-violet – almost black-color of the leaves. These cultivars are the sources
of the new valued biological characteristics for the breeding work to create the perspective
domestic cultivars in the conditions of the south of Russia.
This study was funded by a research grant No. 14-50-00079 of the Russian Science
Foundation.
Keywords: garden roses, lilac, tulip, iris, daylily, chrysanthemum, clematis, canna.
113
113
Расширение биологического разнообразия коллекций цветочно-
декоративных растений Никитского ботанического сада
В.К. Зыкова, З.К. Клименко, Н.В. Зубкова, Л.М. Александрова, И.В. Улановская,
С.А. Плугатарь, Н.В. Смыкова, И.В. Кравченко
ФГБУН «Ордена Трудового Красного Знамени Никитский ботанический сад –
Национальный научный центр РАН», Никитский спуск 52, пгт Никита, г. Ялта,
298648, Российская Федерация; e-mail: zykova.vk@mail.ru
114 114
Постерные презентазии
Poster Presentations
115
115
116 116
The role of a number of culture medium components in obtaining viable
regenerating plants of alfalfa
Sadagat Asadova
Institute of Molecular, Biology and Biotechnology, Matbuat avenue, 2A, Baku, Azerbaijan;
E-mail: biotexnoloqaz@mail.ru
It is known that with the use of certain methodological approaches, most plants can be
regenerated from cell tissue. But not always these techniques are quite effective, since the
reaction of different plant species to similar chemical and physical conditions of cultivation
is not the same. Regulation of the expression of certain genes by metabolic processes is an
undeniable fact, but direction and nature of metabolism in in vitro conditions has its own
specifics. If micropropagation involves the production of regenerating plants via indirect
morphogenesis, then this process consists of three stages: the induction of callusogenesis,
organogenesis or somatic embryogenesis, the production of regenerating plants. At each of
the three stages the mineral and hormonal composition in the culture medium varies to a
certain extent. Until now it is unclear what role one or another component of the culture
medium used at the stage of initiation of cell culture can play in the production of normal
regenerating plants. Moreover, interactions between mineral and hormonal components
can take place. And the question of how these interactions can influence the whole process
of micropropagation still remains undetermined. Therefore, with the use of the method of
mathematical experiment planning in the example of alfalfa, we tried to trace the degree of
participation of a number of culture medium components in the process of obtaining
normal viable regenerating plants.
A regular plan for fractional factorial analysis 210-5 (1/32 complete factorial analysis
210) was adopted for the planning of the experiment. The number of normal regenerating
plants obtained on the five arbitrarily taken embryoids was taken as optimization
parameters. Based on the obtained data 3 regression equations, optimum for
callusogenesis, embryogenesis and regenerative plants were calculated.
117
117
The genetic resources database of fruit plants in Mountainous Dagestan
as a basis for development of adapted cultivars
The results of a longtime (20 years) study of natural populations and local cultivars of
apricot, apple, pear, plum, peach and grapes in Mountainous Dagestan are presented. Their
diversity (361 cultivars and forms) are explained by the intensity of microevolutional
processes and the duration of civilization. The experimental plot s collection of clones and
selectional hybrids of studied fruit crops is developed and become more complete. A
number of limitations are noted in the forms and cultivars available in the collection: early
flowering of the apricot which for many years results to the loss of the greater part of the
crop because of recurrent frosts, lethality by rusty spotting and scab of pear and apple
cultivars and forms, wilt (Monilia) and shot-hole disease (Clasterosporium carpophilum) for
apricot and peach cultivarss. The developed selection program for creating its own adapted
apple tree assortment (87 hybrid forms) and apricot (22 hybrid forms) with the
involvement of the gene pool of Central Asian apricots with genetically fixed later flowering
is presented. The prospect of new direction in horticulture is grounded. It is intended to
enrich the gardens with cultivars containing a large number of biologically active
substances and antioxidants. The advantage of local cultivars and natural forms in the
significant content of these substances is shown in comparison with the introduced
cultivars that are widespread in the region. Methods for optimizing the location and
acceleration of fruiting have been developed and introduced in testing of large number of
hybrids’ seed progeny, material from natural population, introductions and grafting
selection forms.
Keywords: genetic resources, fruit crops, population forms, local cultivars, collections,
Mountainous Dagestan, adapted cultivars.
118 118
База данных генетических ресурсов природных популяций и
местных сортов плодовых растений Горного Дагестана как основа
для создания адаптированных сортов
119
119
Screening hybrids seedling of Iris × germanica L. to salinity
In order to identify the salt tolerance of hybrids an experiment was conducted with four
different hybrids of German Iris under different levels of salinity tension with
concentrations of 4, 6, 8 and 12 ds/m from a source of sodium chloride. This experiment
was factorial based on randomized complete blocks design (RCB) with three replications in
the research park of Mahallat municipality. The result showed that by physiological and
morphological factors, the hybrids with code=23 were better than other hybrids, and the
less level of morphological factors was observed in the hybrids with code=14; the less rate
of physiological factors was identified in the hybrids with code=18, and the highest level of
proline was specified in the hybrids with code=14. The result showed that the highest level
of peroxidase was at the salinity level of 8 ds/m; the highest level of chlorophyll (a, b) and
carotenoid were noted at salinity level of 6 ds/m and the highest level of proline was
observed at salinity level of 12 ds/m. The interaction result showed that the highest level of
chlorophyll (a, b) was evaluated in the hybrid with code=54 at the salinity level of 12 ds/m.
Also the highest level of carotenoid at the salinity level of 6 ds/m was identified in the
hybrid with code=54; the highest level of proline at the salinity level of 4 ds/m was
observed in the hybrid with code=54. The different levels of salinity make it is possible to
screen different hybrids and special selection hybrids for a specific salinity level, and we
consider this method as a rather promising one.
120 120
Biologically active compounds in Myrtus communis L.
Gardens – National Scientific Center of the RAS", Yalta, 298648, Russian Federation;
E-mail: tkdizain@yandex.ru
2Pyatigorsk Medical-Pharmaceutical Institute-Branch of the State Public Health Service of
the Volga Federal Medical University of the Russian Ministry of Health, 11 Kalinin Prospect,
Pyatigorsk, Stavropol Krai, 357532, Russian Federation
The common myrtle (Myrtus communis L.) is a promising medicinal plant for the creation
of herbal medicines with anti-inflammatory, antidiabetic, anti-mutagenic, proapoptotic,
antiatherogenic, antiulcer, insecticidal and protozoicidal properties. Thus, the dynamics of
polyphenolic compounds and myrtle essential oil extraction depending on the ethyl
alcohol volume fraction in the extractant (water-ethanol mixture) was studied in the
samples grown under the conditions of the Southern coast of Crimea. The total phenolics
content was determined according to the Folin-Ciocalteu method using Uv-Vis
spectrophotometry. Investigations of the phenolic complex qualitative composition was
carried out using the HPLC/DAD method. Component composition of the aromatic
substances in the aqueous ethanol extracts was found out using the GC/MS method. The
maximum of total phenolics was revealed in 70% water-ethanol extract. The analysis of
the phenolic compounds qualitative composition demonstrated that in aqueous and 20%
water-ethanol extracts, the maximum amount of gallic acid (up to 708 mg/dm3) was
presented. An increase in the ethanol volume fraction in the extractant to 90% led to
decrease in the gallic acid concentration to 25.8 mg/dm3. Ethanol volume fraction in the
extractant has a slight effect on the ellagic acid extraction – from 21.9 to 42.3 mg/dm3.
When the ethanol volume fraction was from 20 to 40% quercetin glycosides were
extracted. The maximum of myricetin-3-O-rhamnoside (1,045.48 mg/dm3) was obtained
when the raw material was extracted with 70% aqueous ethanol. The studies of the
volatile substances qualitative composition in the water-ethanol extracts demonstrated
that the main components of myrtle essential oil (myrtenyl acetate, linalool and
eucalyptol) were identified only in 70% and 90% aqueous ethanol extracts. Thus, 70%
water-ethanol extract of the myrtle raw material had the maximum concentrations of total
phenolics, gallic acid and myricetin-3-O-rhamnoside, as well as all the main myrtle volatile
compounds (myrtenyl acetate, linalool and eucalyptol) were also found in the extract. This
extract is a promising source of biologically active substances.
121
121
Биологически активные соединения Myrtus communis L.
Национальный научный центр РАН», Никитский спуск 52, пгт Никита, г. Ялта,
298648, Российская Федерация; e-mail: tkdizain@yandex.ru
2Пятигорский медико-фармацевтический институт-филиал ФГБОУ ВО ВолгГМУ
122 122
Clonal propagation of grapes in vitro and adaptation ex vitro
To improve the plants today different biotechnology methods are used. Micropropagated
in vitro plants are not directly transfer to environmental conditions and need a period of
adaptation.
The objects were complex-resistant grape cultivars. As an initial material, intensively
growing green shoots of grapes were taken, which were cut into single-eyed cuttings and
further carried out the isolation of meristems in laminar flow cabinet. The experiments
included cultivars ‘Augustin’ and ‘Moldova’.
Growth regulators are one of the most important and indispensable components of
the culture medium. Their careful selection and identification of optimal concentrations
make it possible to increase the effectiveness of grape micropropagation method. The
experiments showed that shoot regeneration from isolated apexes occurred at all
concentrations of 6-BAP. More effective was 6-BAP at 0.5-1.0 mg L-1. To accelerate the
process of microshoots elongation the effect of gibberellic acid (GA3) in various
concentrations and combination with 6-BAP was investigated. The combination of 0.5 mg
L-1 6-BAP and 1.0 mg L-1 GA3 the best result was shown. This combination of growth
regulators accelerated the elongation of microshoots, and in two weeks the size of shoots
reached 25-26 mm.
At the stage of ex vitro plant adaptation the effect of lignohumate was studied due to a
number of its positive properties, first of all, the ability to increase the resistance of plants
to unfavorable environmental factors. By the soluation of this drug roots were treated, and
also after planting the plants immediately watered the substrate. For optimal effect
establishing the, different concentrations of the drug were studied. As a control the distilled
water was used. Lignohumate at all concentrations had a positive effect on plants. It is
especially necessary to note the positive effect of lignohumate on the leaf area increase.
The height of the plant was increased after 30 day adaptation with 1.0 g L -1 drug
concentration.
123
123
Клональное размножение винограда in vitro и адаптация ex vitro
Российская Федерация;
2Комплексный научно-исследовательский институт РАН, г. Грозный, Российская
124 124
Biochemical composition changes in Hydrangea macrophylla leaves
during vegetative period
The gas chromatography-mass spectrometry method was used to study ethanol extracts of
Hydrangea macrophylla Ser. leaves. As a result, 11 endogenous organic substances were
identified and extracted: isovaniline, vanillic acid, ethyl ester, ethyl carbohydrate, α-d-
glucopyranoside, palmitic acid, palmitic acid ester, acyclic monounsaturated diterpene,
alcohol - phytol, octadecyl and aromatic hydrocarbon from the group of diarylethylenes
2,4'-Dihydroxy-stilbene, squalene, vitamin E, phytosterols γ-sitosterol / β-sitosterol. The
most interesting among them are antioxidants, in particular aromatic hydrocarbon from
the group of diarylethylenes (2,4'-dihydroxy-stilbene), squalene, phytosterols γ-sitosterol /
β-sitosterol. During the vegetative period from March to November, changing in the
content of these compounds in plant leaves was observed. In the most favorable period
with optimal hydrothermal conditions at the beginning of vegetation (March-April)
maximum concentration in the leaves was noted (2,4'-Dihydroxy-stilbene - 14.0%, γ-
sitosterol / β-sitosterol - 5.7%). During this period, there was no squalene observed.
However, during the flowering period coinciding with unfavorable hydrothermal
conditions for H. macrophylla in the region, the content of 2,4'-dihydroxy-stilbene and γ-
sitosterol / β-sitosterols decreased to 8.5% and 1.7%, respectively. In the unfavorable
conditions Squalene was found in all H. macrophylla cultivars, its content was 2.4% in
average. Cultivar effect is important on the intensity of metabolic processes in H.
macrophylla plants. So relatively resistant cultivars ('Altona', 'Draps Wonder', 'Admiration')
were characterized by the greatest content of 2,4'-Dihydroxy-stilbene (10.6%, 10.8% and
15.6%, respectively ) and γ-sitosterol / β-sitosterol (2.0%, 1.9% and 2.1%, respectively)
during the period of intensive vegetation. On the other hand, drought susceptible cultivars
'Madame Faustin' and 'Harlequin' showed the lowest content of these substances (5.1%
and 1.6%) and (1.1% and 1.4%), respectively.
125
125
Динамика биохимического состава листьев Hydrangea macrophylla в
течение вегетации
126 126
Intraspecific polymorphism of Linum usitatissimum L. by
morphophysiological parameters of seeds
To provide a growing people population with organic food and in the context of a climate
change, the expansion of the range of cultivated plants is gaining increased attention. The
culture of flax, which has become widespread in the Republic of Belarus, remains poorly
studied in the agricultural zone of the Tyumen region.
Successful introduction is determined by the high ecological and biological potential of the
seeds. The aim of the research is to search morphophysiological criteria of seeds for rapid
testing during the introduction of flax.
Seeds of 17 cultivars of flax grown on cultivated sod-podzolic soil (humus – 3.67, pH -
6.6) of the experimental biostation Tyumen State University "Lake Kuchak" (Tyumen
region, Nizhnetavdinsky region) in germination energy and laboratory germination
corresponded to the State standard (GOST) and not inferior to the seeds of the Belarusian
reproduction. In comparison, the average population values of studied reproductions by
the morphometric parameters (length, width) and the shape of the seeds, length of shoots
and roots of the seedlings, no statistically significant differences were found. In structure of
raw biomass of seedlings from the seeds of both reproductions, the proportion of shoots
has been predominated (66.6-62.4%). At the same time, sprouts of cultivars ‘Vesta’,
‘Velizhsky Kriazh’, ‘Fliz’ and ‘Biruza’ from the seeds of Tyumen reproductions formed more
powerful primary root system (44.5-64.3%).
To understand the mechanism of adaptation of flax in northern agrocenoses and to
study the physiological status of seeds, was used optical counter of chlorophyll Spad 502
(Minolta Camera Co., Ltd., Tokyo, Japan) which provides express diagnostics without
seizure of plants. The Spad measurement is based on the optical density calculated from
the absorbance of leaf at 650 nm (maximum absorption of chlorophylls a and b) and optical
density at 940 nm, taking into account the thickness of the leaf. The informative nature of
the application of this criterion in the selection of adaptive forms of flax is confirmed by the
positive correlation with the dimensions (length, width) and the mass of 1000 seeds (r =
0.20, 0.65, 0.49), root length (r = 0.71), linear the size of the leaves (r = 0.16, 0.49).
The selection of forms with valuable characteristics can be carried out at the initial
stages of ontogenesis in simulated conditions. In this case, an objective evaluation is
achieved on basis of a complex of characteristics of the biological status of seeds and the
morphophysiological properties of seedlings.
127
127
Внутривидовой полиморфизм Linum usitatissimum L. по
морфофизиологическим показателям семян
128 128
Development of large-scale ecological-phytocoenotic maps as an
informational basis for regional plant diversity monitoring and
conservation
Modern geobotanical maps give a synthetic knowledge about the regional flora and
vegetation. They play an important role for fundamental and applied studies of nature
conservation and biological resources. The main aim of the study is development of the
large-scale vegetation map of steppe biome in Crimea peninsula for the assessment of
ecological peculiarities of biotopes, phytocoenology and various parameters of plant
species distribution. The basis for the study was 340 relevés of steppe communities
collected from Kazantip peninsula (Eastern Crimea). The classification of plant
communities was carried out using the Braun-Blanquet approach. The plot records were
classified by TWINSPAN. Syntaxa ecology was studied with DCA ordination. The spatial
structure of steppe vegetation was studied using satellite image WorldView-2 (resolution
1.8 m). The total diversity of steppe biome plant communities was assigned to five classes
14 orders, 16 alliances and 26 associations. Results of DCA ordination demonstrated an
arrangement of vegetation associations along leading ecological factors. The Axis 1 was
interpreted as complex gradient of two factors – soil salinity and soil moisture regime. Axis
2 of the DCA was explained in terms of substrate type (petrophytic factor). Four ecological
sets of syntax observed along leading axes represent micro-combinations of plant
communities related to certain landforms in the landscape. These micro-combinations
were used as basic spatial categories for the map legend construction. Application of
satellite images of high resolution (1.8 m) allowed developing the large-scale vegetation
map representing tree levels of spatial vegetation structure of halophytic-steppe vegetation
mosaic – certain plant communities, micro- and meso-combinations of vegetation types.
The map was used for evaluation of ecology and distributions of Red Data Book plant
species in their biotopes and for monitoring.
This study was funded by research grant No. 14-50-00079 of the Russian Science Foundation.
Keywords: vegetation mapping, biodiversity, rare species, plant ecology, steppe, Crimea.
129
129
Создание крупномасштабных эколого-фитоценотических карт как
информационная основа мониторинга и сохранения регионального
фиторазнобразия
130 130
Drought resistance and photoactivity of leaf apparatus of peach cultivars
under conditions of the Southern Coast of Crimea
Local climate is highly suitable for the cultivation of various fruits including peach (Prunus
persica (L.) Batsch). Peach fruits are in great demand all over the world. The main limiting
factor for the peach growing in the Crimea is the moisture deficit during the setting of
reproductive buds and the ripening time. Therefore, the study on the mechanism for
adapting the peach leaf apparatus to dehydratation is a very important challenge under
conditions of the Southern Coast of Crimea.
Investigations were carried out in the Nikita Botanical Gardens. The project
comprised six cultivars of foreign origin. The cultivar ‘Cardinal’ has been used as control.
The weather conditions during the experimental period were extremely unfavorable for
the plants. This fact made it possible to evaluate objectively the introduced peach cultivars
for their adaptivity to the drought conditions (at 30.2°С on average and 37.0°С maximally
and under precipitation of 0 mm). The water content in leaves, the water loss over the
course of wilting and the turgor restoration ability have been measured according to
conventional techniques (Lishchuk et al., 1991; 1999). The photosynthetic activity has been
evaluated according to indicators of chlorophyll fluorescence induction (Kautsky effect)
determined by means of the portable fluorimeter "Floratest". It has been ascertained that
studied peaches were mainly susceptible to drought under stress conditions of August. The
maximum turgor restoration was shown by cultivars ‘Asmik’ (80%) and ‘Zempush’ (68%),
the minimum one – by ‘Favorita’ (22%). The use of parameters of photo-induced curve of
chlorophyll fluorescence (Fm; (Fm-Fst)/Fm; Fm/Fst) allowed to make an objective assessment
of introduced peach cultivars relating to their resistance to the summer drought and to
determine the varietal differences. The cultivars ‘Asmik’ and ‘Zempush’ have kept the most
sustainable photosynthetic activity.
This study was funded by the research grant No. 14-50-00079 of the Russian Science
Foundation.
131
131
Устойчивость к засухе и фотоактивность листового аппарата сортов
персика в условиях Южного берега Крыма
132 132
Effectiveness of high cytokinin levels on strelitzia micropropagation
Junia R Mendonça Figueiredo1, Patrícia Duarte de Oliveira Paiva2, Diogo Pedrosa Corrêa
da Silva2, Renato Paiva3, Rafaela Ribeiro de Souza3, and Michele Valquíria dos Reis3
1Universidade Vale do Rio Verde UNINCOR, Três Coraçes-MG, Brazil
2Universidade Federal de Lavras UFLA, Departamento de Agricultura, 37200-000 Lavras-
Gerais, Brazil
The in vitro propagation of strelitzia (Strelitzia reginae Banks) seedlings may have
limitations during multiplication, and alternatives to optimize this process are needed
which may contribute to improve the propagation of many other species. In this way, the
objective was to optimize the multiplication process of strelitzia by modifying the cytokinin
concentrations, using light levels of these compounds. Shoots with 2.0 cm long obtained
from in vitro embryo germination were excised and inoculated in MS medium
supplemented with 0, 5, 10 and 20 μM 6-benzylaminopurine (BAP) or 0, 5, 10 and 15 μM
thidiazuron (TDZ) and maintained for 60 days in a growth room. The results showed that
20 μM BAP provided a better shoot formation, with an average of 1.46 shoots per explant,
being observed in some explants up to 4.0 shoots which was 46% higher as compared with
the control. Besides the increase in shoot number, it was observed a reduction in shoot
length (2.71 cm), as well as an increase in oxidation of the culture medium. Otherwise, the
use of TDZ was not effective to stimulate the formation of new shoots. The results showed
that 20 μM BAP can be used for in vitro multiplication of streliztia.
The authors are thankful to CNPq, Capes and Fapemig for the financial support.
133
133
Variation of fruits morphometric parameters of date plum (Diospyros
lotus L.) germplasm collection
Olga Grygorieva1, Svitlana Klymenko1, Yulia Vinogradova2, Olena Vergun1, Jan Brindza3
1M.M. Gryshko National Botanical Gardens of Ukraine, National Academy of Sciences,
134 134
Influence of a soil drought on photosynthesis and a water regime of
Viburnum tinus L. in the conditions of the Southern Coast of the Crimea
135
135
Влияние почвенной засухи на фотосинтез и водный режим
Viburnum tinus L. в условиях Южного берега Крыма
136 136
In silico identification of genes of antimicrobial peptides in genome of
Solanum lycopersicum str. Heinz 1706
This work was supported by the Russian Science Foundation (grant No. 16-16-00032).
137
137
In silico идентификация генов антимикробных пептидов в геноме
Solanum lycopersicum str. Heinz 1706
Е.А. Истомина1, А.С. Ковтун2, А.С. Касьянов1, Т.В. Коростылева1, Т.И. Одинцова1
1Институт общей генетики им. Н.И. Вавилова Российской академии наук, ул. Губкина
138 138
Structural and biochemical characteristics of clematis plants in vitro
Natalya Ivanova, Valentina Brailko, Oksana Grebennikova, Anfisa Paliy, Irina Mitrofanova
Federal State Funded Institution of Science "The Labor Red Banner Order Nikita Botanical
Gardens – National Scientific Center of the RAS", Yalta, 298648, Russian Federation;
E-mail: invitro_plant@mail.ru
The world assortment of clematis includes more than 3,000 cultivars. Using biotechnology
methods give possibility to obtain virus-free valuable and promising clematis cultivars for
subsequent in vitro propagation and preservation. The objective of this study was to reveal
the morphological-anatomical and biochemical characteristics of four in vitro improved
clematis cultivars: 'Nikitsky Rozovy', 'Alpinist', 'Crystal Fountain' and 'Madame Julia
Correvon'. Isolated sterile vegetative buds were cultured on MS medium with 0.5-0.7 mg L-
1 BAP and 0.01-0.03 mg L-1 NAA. 3.8±0.7 normally developed microshoots (up to 3.0-3.5 cm
long with 4-5 internodes) per explant in 'Nikitsky Rozovy', 3.7±0.5 – in 'Crystal Fountain'
and 2.5±0.4 – in 'Madame Julia Correvon' were obtained. The shape of the leaf blades
retained varietal characteristics of the initial mother plants. It was determined that the leaf
size was a more cultivar-specific characteristic. Leaves were thin from 83 to 182 µm,
hypostomatic, with a differentiated 4-5-layer mesophyll (palisade index 0.29-0.37).
Vascular bundles were closed collateral, stomatal apparatus were numerous (up to 136
stomata/mm2, stoma pore length 28-51 μm), of anomocyte type, trichomes were simple
one- or two-celled (up to 143 μm long, from 2 to 16 trichomes per 1 mm2). Epidermal cells
were sinuous of irregular amoeboid shape. The assessment of assimilation processes
demonstrated partial photoinhibition. The content of the protective compounds in all
clematis cultivars was quite high: proline concentration varied from 8.63 μg g-1 ('Alpinist')
to 31.00 μg g-1 ('Crystal Fountain'), ascorbic acid – from 30.10 mg/100g ('Nikitsky Rozovy')
to 35.64 mg/100 g ('Crystal Fountain'), phenolic compounds – from 559 mg/100 g
('Nikitsky Rozovy') to 1026 mg/100 g ('Crystal Fountain'). Enzyme activity was low:
catalase - 14.27-18.28 gO2/g·min, superoxide dismutase – 19.52-25.45 units/g, polyphenol
oxidase – 0.078-0.109 units/g·s. The minimum activity of enzymes was noted in the
cultivar 'Crystal Fountain' and the maximum – in 'Nikitsky Rozovy'.
This study was funded by the research grant Nо. 14-50-00079 of the Russian Science
Foundation.
Keywords: Clematis L., microshoot, leaf blade structure, photosynthetic activity, protector
compounds, enzyme activity.
139
139
Структурные и биохимические характеристики растений клематиса
в культуре in vitro
Н.Н. Иванова, В.А. Браилко, О.А. Гребенникова, А.Е. Палий, И.В. Митрофанова
ФГБУН «Ордена Трудового Красного Знамени Никитский ботанический сад –
Национальный научный центр РАН», Никитский спуск 52, пгт Никита, г. Ялта,
298648, Российская Федерация; e-mail: invitro_plant@mail.ru
140 140
Adaptive potential of the walnut gene fund
Abiotic stressors cause significant economic damage to horticulture and are one of the
main causes that negatively affect the productivity of crops. Along with social and economic
problems, the efficiency of industrial fruit growing in the Russian Federation depends to a
large extent on the low adaptive potential of cultivated cultivars. Without the creation of
new integrated systems that increase the production of domestic fruit products, it is
difficult to solve the task of replacing fruit imports. In such conditions, the use of the most
effective methods for evaluation the genetic potential of the initial forms and hybrid
material will allow to increase the productivity of the selection process and to obtain the
new generation of highly productive and adaptively resistant cultivars of walnut. In the
course of the research, 27 walnut cultivars from the gene fund collection in the Nikita
Botanical Gardens were compared in 10 main economic characteristics with the developed
ideal model of the cultivar: the period of entry into fruiting (4 years), fruit size (3
points), fruit mass (26 g) , kernel taste (5 points), kernel output (60%), fat and protein
content (70% and 22%, respectively), productivity (50 kg/t), frost resistance (3 points),
drought resistance (32 points), heat resistance (3 points). Cluster analysis of the
experimental data was carried out using the computer program Statistics 10. The
distribution of cultivars in 4 groups was carried out using the calculated integral criteria of
all determined indicators that characterize their degree of similarity with the ideal model: I
– ‘Sladkoyaderny’, ‘Gurzufsky’, ‘Doliny’, ‘Skabery’, ‘Partizansky’, ‘Yuzhnoberezhny’,
‘Pozdnetsvetushchy’, ‘Arkad’ and ‘Bubenchik’; II – ‘Bospor’, ‘Starokrymsky’, ‘Kacha’, ‘Pioner
Kryma’, ‘Originalny, Zhemchuzhny’, ‘Belbeksky Ranny’, ‘Krymsky Skoroplodny’ and
‘Uigursky’; III – ‘Alminsky’, ‘Burlyuk’, ‘Konkursny’ and ‘Pamyaty Pasenkova’; IV –
‘Moldovskaya Bomba’, ‘Podarok Valentiny’, ‘Bomba Chkalovskaya’ and ‘Purpurovy’. The
following cultivars proved to be the closest to the model of the cultivar (18 units): ‘Karlik 3’
(16 units), ‘Purpurovy’ (11 units) and ‘Moldavskay Bomba’ (10 units).
The work was supported by the grant of the Russian Science Foundation No. 14-5-00079.
Keywords: walnut, varieties, gene fund collection, variety model, cluster analysis.
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141
Адаптивный потенциал генофонда ореха грецкого
142 142
Fluorescence intensity of chlorophyll in Chaenomeles leaves in
connection with drought resistance
One of the factors influencing the productivity of chaеnomeles is the resistance to the
effects of water stress. Conditions of prolonged soil and air drought negatively affect the
state of plants and the crop quality, as cause irreversible violations to the functional state
of the photosynthetic apparatus. To identify genotypes with high potential drought
resistance, the study was made of the chlorophyll photoactivity parameters under different
conditions of the water regime using a portable fluorometer "Florаtest" in five
сhaеnomeles breeding forms, originated from Chaenomeles japonica, C. speciosa,
C. cathayensis, C. × superba.
The initial level of fluorescence after irradiation (Fo), maximum (Fm) and stationary
(Fst) after the light adaptation were recorded. It was found that for genotypes of all
Chaenomeles species, a rapid loss of water in wilting conditions (from 36 to 93%) and a
significant restoration of the leaf turgor (from 38 to 80%) are typical. Saturation of leaves
with water led to an increase in the fluorescence parameters for all assessments. After
significant dehydration, the maximum level of fluorescence (Fm) and variable fluorescence
(Fv = Fm - Fo) decreased from 17 to 51% in most genotypes. In PX-3-12, on the contrary,
these parameters were elevated, and the level of water loss by leaves was the lowest. The
parameters of photosynthetic activity (FA = (Fm - Fst) / Fm) and viability (Fm / Fst) were
increased in PX-3-12 and XK-2 from 7 to 21%. The same genotypes, as well as РХ-5-19,
were characterized by the least variability of all fluorescence parameters under different
water availability regimes, which indicates their increased adaptive capacity.
This study was funded by the research grant No. 14-50-00079 of the Russian Science
Foundation.
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143
Интенсивность флуоресценции хлорофилла в листьях хеномелеса в
связи с засухоустойчивостью
144 144
The content of phenolic compounds in ornamental peach flowers
The aim of the investigation was to study the qualitative composition and content of
phenolic substances in flowers of ornamental peach genotypes with different taxonomic
origin. The sum of phenolic compounds was determined using Uv-Vis spectrophotometry.
Phenolic acids and flavonoids were identified and quantified by the HPLC/DAD method.
A high total content of phenolic compounds during the budding and mass flowering of
five ornamental peach genotypes originated from Prunus mira, P. davidiana, P. kansuensis,
P. persica (20.3-46.9 mg·g-1 d.w.) was found. The maximum content of phenolic compounds
was determined in the buttons of cultivar 'Malenky Princ' (46.9 mg·g-1 d.w.) and cultivar
'Zhiselle' (46.8 mg·g-1 d.w.) and in the flowers of 'Malenky Prince' (37.3 mg·g-1 d.w.).
Phenolic compounds of peach buttons and flowers are represented by flavonoids and
hydroxycinnamic acids, among which are identified neochlorogenic, chlorogenic, p-
coumaric acids, rutin and kaempferol. The 'Malenky Princ' buttons was characterized by
the highest concentration of neochlorogenic acid (3.6 mg·g-1 d.w.) and flowers of this
cultivar contained the maximum value of the sum of dicofoelchinic acids (1.7 mg·g-1 d.w.).
The highest concentrations of chlorogenic (3.5 mg·g-1 d.w.) and p-coumaric acids (0.6 mg·g-
1 d.w.) were detected in buttons and flowers of 'Zhiselle', respectively. The flowers of
cultivar 'Belosnezhka' were distinguished by the content of routine (1.0 mg·g-1 d.w.) and
kaempferol (0.5 mg·g-1 d.w.).
The results show that buttons and flowers of ornamental peach can be valuable
sources of phenolic antioxidants. Individual selection will be promising in breeding for high
content of phenolic compounds in flowers.
This study was funded by the research grant No 14-50-00079 of the Russian Science
Foundation.
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145
Содержание фенольных соединений в цветках декоративного
персика
Ключевые слова: Prunus sp., сорт, гибрид, флавоноиды, фенольные кислоты, цветки,
бутоны.
146 146
Cold storage of in vitro apple germplasm in Kazakhstan
Kazakhstan
2Institute of Plant Biology and Biotechnology, Timiryazev Str.,45, Almaty, 050040,
The duration of in vitro cold storage (+3оС and +4оС, 7-15 μmol m-2s-1 light intensity and 10
h photoperiod) in plastic breathable packages of three apple cultivars ‘Voskhod’, ‘Golden
Delicious’ and ‘Maksat’ were studied. The effect of three sources of carbohydrates (3%
sucrose, 2% or 3% mannitol, or 2% sucrose + 2% mannitol) in MS medium with 0.5 mg L -1
BAP and 0.1 mg L-1 IBA or without growth regulators (PGRs), the effect of abscisic acid (AA)
in three concentrations (0.1, 0.5 and 1.0 mg L-1) and three concentrations of nitrogen (N03)
(100%, 50% and 25%) were determined.
Apple shoots remained viable for 18-20 months without repeated multiplication
under conditions of cold storage on a standard (control) MS medium with 3% sucrose
(1962) with and without PGRs. The effect of the cultivar and medium composition on the
duration of cold storage was determined. The increase in storage time was with a decrease
in the nitrogen concentration in the MS medium to 25% or 50%. In these cases, the storage
time was extended to 31 months without PGRs and up to 28 months with PGRs compared
to controls. The shoots depending on the cultivar remained viable (rating ≥ 2) from 23 to
25 months on MS medium and a combination of carbohydrates of 2% sucrose + 2%
mannitol regardless of the hormonal background. The addition of abscisic acid regardless
of the concentration in the medium with 3% sucrose increased storage also up to 25
months. The effect of the cultivar was insignificant. The longest storage time was recorded
for ‘Golden Delicious’ cultivar (31 months), the shortest storage time for ‘Maksat’ cultivar
(28 months).
In vitro germplasm bank contains 27 apple cultivars on an optimal MS medium with
50% N03 and 2% sucrose + 2% mannitol without PGRs.
This study was funded by Ministry of Education and Science of the Republic of Kazakhstan,
project No. AP05132995.
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147
In vitro establishment of grape explants in active plant growth and
dormancy periods
The aim of the research was to study and compare morphogenetic capacity of grape
explants (cultivars ‘Regent’, ‘Marquette’, ‘Ventura’, ‘Zilga’, ‘Agat donskoy’ and ‘Aleshenkin’)
during dormancy and active growth periods at the initiation stage of in vitro culture in
conditions of the Republic of Belarus. Previously, the plants have been tested for the main
sap-transmittable viruses of grape: GFLV, GLRaV-1, GLRaV-2, GLRaV-3, GVA and GFkV. The
explants were buds without cover scales from hardwood cuttings in dormancy period; and
buds without cover scales from softwood cuttings in period of active growth. The
sterilization scheme: main sterilizing agent – 30% hydrogen peroxide; in active growth
period, the exposure was 10 min, in dormancy period – 15 min. The initiation medium was
modified Murashige and Skoog medium with 1.1 mg L-1, 6-BA and 0.09 mg L-1 NAA. During
the study of explant morphophysiological capacity of grape cultivars at dormancy period, it
was established that plant genotype with high degree of reliability had determined their
ability to regeneration processes in in vitro culture (p <0.001). Cultivars ‘Agat donskoy’ and
‘Regent’ have maximum capacity in selected conditions (100%). Cultivars ‘Zilga’ and
‘Ventura’ have maximum percent of not developed explant: 69.5% and 65.0%, respectively.
In addition, callus was actively formed at the base of 41.7% of explants of cultivar
‘Marquette’. Highly reliable influence of period of initiation on regeneration processes of
explants of studied cultivars ‘Agat donskoy’, ‘Regent’ and ‘Marquette’ was detected (p
<0.001). Activation of regeneration processes of explants on studied medium was noted
only in dormancy period. In dormancy period, the number of explants which have
developed in plants was 72.2%, which is 1.9 times higher than in period of active growth.
The number of undeveloped explants was not more than 13.9%.
This study was funded by the Belarusian Republican Foundation for Fundamental Research,
grant No. B17-097.
148 148
Введение в культуру in vitro эксплантов рода Vitis L. в период
активного роста растений и в период покоя
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149
Phytochemical analysis of the vegetative organs of hybridogenous
complex Reynoutria Houtt.
N.V. Tsitsin Russian Academy of Sciences, Botanicheskay str. 4, Moscow, 127276, Russian
Federation; E-mail: alla_gbsad@mail.ru
2Federal State Budgetary Scientific Institution “All-Russian Scientific Research Institute of
Medicinal and Aromatic Plants”, Grina str. 7, Moscow, 117216, Russian Federation
Keywords: Reynoutria, leaf, build-up cone, flavonoids, ascorbic acid, organic acids.
150 150
Фитохимическое изучение вегетативных органов гибридогенного
комплекса Reynoutria Houtt.
ботанический сад имени Н.В. Цицина Российской академии наук, ул. Ботаническая,
д.4, г. Москва,127276, Российская Федерация; e-mail: alla_gbsad@mail.ru
2Федеральное государственное бюджетное научное учреждение “Всероссийский
151
151
The regulatory and legal framework for creating virus-free planting
material of horticultural crops
152 152
Нормативно-правовая база создания безвирусного посадочного
материала садовых культур
153
153
Experimental variability of grapes in Vitaceae family based on the use of
allotetraploidy and in vitro embryo culture
Formation of grapes samples occurred in the course of natural evolution and is possible in
the process of experimental evolution under artificial generative hybridization. Crossing
forms of grapes with a different number of chromosomes suggest the need to combine
several methods that take into account the physiological characteristics of the initial forms.
An attempt was made to achieve intergeneric hybrids Vitis Ampelopsis and
Vitis Parthenocissus within the Vitaceae family using tetraploid genotypes of the initial
forms. In order to obtain tetraploid seedlings (diploid gametes), buds broken from
dormancy and flower clusters were treated with 0.5%, 1% and 2% colchicine for 5-14
days before meiosis. The intergeneric incompatibility was circumvented by growing
seedlings from in vitro cultured embryos isolated from immature seeds for 40 days after
pollination. An intermediate dissection pattern of the leaf typical of the species of the
genera Vitis and Ampelopsis used in intergeneric hybridization by the allotetraploidy
method was registered in a number of hybrid seedlings. More than that, this is a proof of
the biologically determined possibility that forms from distant crosses may have emerged
in nature in the course of natural evolution. Due to the fact that it was more successful to
obtain plants from the hybridization of Vitis vinifera cultivars of the genus Vitis with the
forms of Ampelopsis genus species, in comparison with the crossing of the Vitis vinifera
cultivars with the forms of the Parthenocissus genus species, can be assume a closer
genetic relationship in the Vitaceae family between the genera Vitis and Ampelopsis, which
are closer in terms of the evolution of the grapes culture as a whole.
154 154
Экспериментальная изменчивость винограда семейства Vitaceae на
основе использования аллотетраплоидии и культуры эмбриоидов
in vitro
155
155
Optimization of potato cultivation process in vitro as influenced by
different spectral composition of LED radiation
Elena Maslova, Vladimir Yatsenko, Yuri Maslakov, Yana Dyachenko, Irina Korsun, Kristina
Degtyareva
Belgorod State University, 85, Pobrdy Str., Belgorod, 308015, Russian Federation;
E-mail: e_maslova@list.ru
Potato is one of the main vegetable crops, for which the most fully technologies of
improvement and clonal micropropagation were developed and an effective culture media
were selected. There are also the number of works on the effect of the light spectral
composition of monochrome lamps on the growth and development of potato plants in
vitro (Kim et al., 2004; Martirosyan et al., 2016; Karimov, 2017). However, the
development of modern optical systems in presently market allows us to produce the
special designs, including polychrome led racks with adjustable spectra both in power and
in composition. All this gives new opportunities to increase the speed and efficiency of
growth and development of potato plants in vitro. The aim of the study was to select
effective methods of potato plants in vitro culture and to assess their growth and
development. As the object of study potato cultivar ‘Udacha’ was selected. The cultivation
of the plants was carried out on modified MS culture medium (Murashige and Skoog,
1962) in separate flat-bottomed tubes. Fluorescent lamps were used as a control. In the
experiment, several variants of the spectral composition of led radiation were tested,
installed for each shelf separately on the phytorack X-bright FitoLed (ELSIS Belgu LLC)
with the help of a programmable controller: 1st variant (the ratio in the spectrum of red,
white and blue) – 57%, 14%, 29%; 2nd variant – 80%, 0%, 20%; 3rd variant – 70%, 0%,
30%; 4th variant – 50%, 18%, 32%. More than 1,000 test tube plants were analyzed. As a
result of 30 days of culture, the following morphometric characteristics of the studied
plants were taken into account: the number of roots formed and their length, length and
thickness of shoots, number of leaves, their width and length, spectrometric
measurements of the chlorophyll pigment concentration by the optical density of the
extract from the cultivated mini-plants were carried out. According to the results of
experiments, it was shown the advantage of growing plants on phytorack and the most
effective modes on the spectral composition of (50%, 18%, 32%) led radiation were
determined.
156 156
Оптимизация процесса выращивания картофеля in vitro под
влиянием различного спектрального состава светодиодного
излучения
Е.В. Маслова, В.М. Яценко, Ю.Н. Маслаков, Я.В. Дяченко, И.С. Корсун, К.А. Дегтярева
Белгородский государственный университет, ул. Победы, 85, г. Белгород, 308015,
Российская Федерация; e-mail: e_maslova@list.ru
157
157
Special features of photosynthetic processes in tissue and organ culture
of horticultural plants
Photosynthesis is the main link for plant organism successful functioning, growth, biomass
accumulation and development. Of particular interest, there are the studies of assimilation
processes in plants cultured in vitro. This is due to the conditionally heterotrophic type of
their nutrition, since carbohydrates are present in the culture medium in all stages of
micropropagation. At the same time, it has been previously established that plant
chlorophyll under in vitro conditions is also capable of photosynthesis. This property is
partly realized due to the low CO2 concentration in culture vessels.
Considering the high sensitivity of photosystem-2 chlorophyll-protein complexes
activity under varying culture conditions, fluorescence induction parameters to estimate
the relative photosynthetic activity (Fm-Fst)/Fm in vegetative organs of some valuable plant
cultivars under in vitro conditions were used. Such plants as Canna × hybrida hort ex
Backer, Chrysanthemum × hortorum Bailey, Clematis L., Rosa × damascena Mill., Lavandula
angustifolia Mill. and L. × intermedia Emeric ex Loiseleur, Diospyros kaki Thunb. and Ficus
carica L. were included in the study. The plantlets were selected under standard culture
conditions in vitro and upon preservation. The assimilating tissues were developed to
various degree. The photosynthetic activity, depending on the genotype and culture
duration, ranged from 0.48 to 0.72 a.u. Active photoinhibition was noted in clematis,
persimmons and lavender cultivars under the culture for more than six months.
During in vitro storage, the indices of chlorophyll fluorescence induction were lower:
from 0.30 to 0.61 a.u. (at the same time in most cultivars (Fm-Fst)/Fm at the level of 0.38-
0.42 a.u.). Positive correlation between the histological characteristics of palisade index
and relative photosynthetic activity was revealed. The rate of the fluorescence decay was
identified as a functional characteristic.
Thus, for a number of horticultural plants, the parameters of relative photosynthetic
activity that characterize the functional state of the plantlets during a certain period of in
vitro culture and preservation were determined. The obtained results can be used to
monitor plant physiological state during the prolonged culture.
This study was funded by research grant No. 14-50-00079 of the Russian Science Foundation.
158 158
Особенности фотосинтетических процессов в культуре органов и
тканей некоторых садовых растений
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159
Micropropagation and preservation of some Crimean flora rare endemic
species from the genus Lagoseris under in vitro culture
Relict endemics of the Mountain Crimea flora are a genetically unique bioecological group.
These include Lagoseris callicephala Juz. and Lagoseris purpurea L. (Asteraceae). Slow
renewal of the studied species, the difficulty of their reproduction, as well as the
intensification of anthropogenic effect resulted in a reduction of their natural areas. In
recent years, for the preservation of plant biodiversity. Biotechnological methods that
ensure both reproduction and conservation of particularly valuable representatives of rare
endemic species, including relict endemics of the Mountain Crimea flora are actively used.
The objective of this work was to develop a micropropagation protocol, to study the
factors affecting morphogenesis in vitro and to introduce Lag. callicephala and Lag.
purpurea explants into the genebank in vitro for further conservation and repatriation to
the natural habitats.
A protocol for in vitro micropropagation has been developed. The effect of plant
growth regulators on the induction of adventitious buds and microrosette development
was revealed. It was found that the combination of BAP (0.1-0.3 mg L-1), IBA (0.1-0.3 mg L-
1) and GA3 (0.1-0.3 mg L-1) was optimal for effective micropropagation of Lag. callicephala
and Lag. purpurea. Rhisogenesis was noted on the hormone-free MS medium and MS
medium supplemented with IBA (0.5-1.0 mg L-1). The optimum physical parameters of in
vitro culture were determined: the light intensity 2,000-2,500 lux and the temperature 22-
23°С. The increase or decrease in light intensity resulted in a decrease of the regeneration
capacity in Lag. callicephala and Lag. purpurea, and it also inhibited plantlet growth. The
greatest number of microrosettes was obtained under the white light. The main factors
affecting the preservation of Lag. callicephala and Lag. purpurea explants in the gene bank
in vitro were determined: the type of optimal explant and its size; mineral composition of
the culture medium; osmotics and retardants concentration; the temperature of deposition
and the time of preservation. Our results demonstrated that growth kinetics changed and
viability of explants was 70-80%.
160 160
Клональное микроразмножение и сохранение некоторых
представителей редких эндемиков флоры Крыма рода Lagoseris в
культуре in vitro
И.В. Митрофанова, О.В. Митрофанова, А.Р. Никифоров, Н.Н. Иванова, Н.П. Лесникова-
Седошенко
ФГБУН «Ордена Трудового Красного Знамени Никитский ботанический сад –
Национальный научный центр РАН», Никитский спуск 52, пгт Никита, г. Ялта,
298648, Российская Федерация; e-mail: nnivanova@yandex.ru
161
161
Aconitum lasiostomum Reichenb. ex Bess. is a promising plant for
biotechnology research
Olga Mitrofanova, Anfisa Paliy, Olga Startseva, Marina Rudenko, Nina Lesnikova-
Sedoshenko, Svetlana Chelombit, Irina Mitrofanova
Federal State Funded Institution of Science “The Labor Red Banner Order Nikita Botanical
Gardens – National Scientific Center of the RAS”, Yalta, 298648, Russian Federation;
E-mail: invitro_plant@mail.ru
162 162
Aconitum lasiostomum Reichenb. ex Bess. – перспективный
растительный объект для биотехнологических исследований
163
163
Ectopic expression of Aseraceae MADS-box genes in Chrysanthemum
morifolium cultivar “Egiptyanka”
Gardens – National Scientific Center of the RAS", Yalta, 298648, Russian Federation; E-mail:
mitiouchkina@rambler.ru
2Branch of Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Pushchino,
Russian Federation
3All-Russia Research Institute of Agriculural Biotechnology, Moscow, Russian Federation
MADS-box genes encode transcription factors playing a key role in floral morphogenesis
control. Different combinations of MADS-proteins are required for floral organ identity and
floral meristem determination. So, modification of its expression patterns results in flower
structure modification and new “unusual” flower shape generation. In this work, we tested
the possibility of chrysanthemum morphology alteration by constitutive expression of
MADS-box genes previously isolated from sunflower and chrysanthemum. The genes
encode two different MADS-activities: CDM37, HAM45 and HAM59 – belong to AGAMOUS
subfamily which specifies stamens and carpel identity, and floral meristem determination;
HAM75 and HAM92 participate in flower induction, determine sepal and petal identity, and
belong to APETALA1 subfamily. Totally 67 independently regenerated plants were
produced by the Agrobacterium-mediated transformation of leaf disks. Transgenes
integration was confirmed by PCR analysis. Constitutive expression of transferred genes
was shown by RT-PCR. Flowering of transgenic chrysanthemum plants was monitored
under controlled greenhouse condition and compared to control non-transgenic plants.
The transgenic plants were analyzed under LD and SD photoperiod. Three types of plant
development alterations were observed: flowering time variation, differences in the stem
height and leaves number, and some inflorescence modifications. Results of the
comparative analysis of transgenic and control plants will be presented.
This study was funded by research grant No. 14-50-00079 of the Russian Science Foundation.
164 164
Эктопическая экспрессия генов MADS-бокса сложноцветных в
хризантеме сорта «Египтянка»
Митюшкина Т.Ю. 1,2, Смыкова Н.В. 1, Шульга О.А. 1,3, Долгов С.В. 1,2
1 ФГБУН «Ордена Трудового Красного Знамени Никитский ботанический сад –
Национальный научный центр РАН», Никитский спуск 52, пгт Никита, г. Ялта,
298648, Российская Федерация; e-mail: mitiouchkina@rambler.ru
2Филиал Института биоорганической химии им. Акад. Шемякина М.М, и
165
165
Regulation of rhizogenic process at clonal micropropagation of
horticultural crops
The relation of the process of root formation to the genotype, medium composition,
substances of phenolic nature, intensity and light spectrum, consequences of the
proliferation stage, ultrasonic and laser exposure and other factors has been established.
According to our data to promote rooting of different actinidia cultivars, it is sufficient to
add 0.25 mg L-1 IBA. In most blackberry cultivars, the best results were obtained on the
culture media supplemented with 0.25-0.5 mg L-1 IBA. The application of IBA in
honeysuckle and arboreal fruit crops rooting was successful only in concentrations of 0.8-
1.0 mg L-1. The application of NAA in horticultural crops, rooting was successful only in
concentrations of 0.1-0.2 mg L-1. For the genotypes with low rhizogenesis capacity in vitro,
such as remote hybrids of intergeneric rowan-pear hybrid, the positive effect was obtained
due to soaking of microcutting bases in the auxin solution with their subsequent planting
in non-hormonal media. For some crops the best rhizogenesis effectiveness is obtained
with the complete carbohydrate concentration in the medium, as well as using alternative
sources of carbohydrate. The concentration of carbohydrate in the medium with 0.1 mol L-
1 per 10-15% increases the rhizogenesis frequency of raspberry microcuttings. The
maximum number of roots of the honeysuckle cultivat ‘Volkhova’ was obtained in the
medium with 0.1 mol L-1 of maltose. Better developed honeysuckle microshoots with the
big diameter of lamina tissue were formed in the sucrose, glucose and fructose
concentrations in the medium with 0.1 mol L-1. No positive effect was marked from the
decrease of carbohydrate concentration in the medium on the process of the rhizogenesis
in the lilac microcuttings. The studies of the rhizogenesis dynamics on the clematis cultivar
‘Zoin Inspiration’ have shown that the maximal speed of rooting in microcuttings occurs
on the media with glucose and fructose. Our attempts to increase the rhizogenesis
effectiveness of microcuttings in vitro by treating them with supersonic and laser
irradiation were successful. In three weeks of cultivation, the rhizogenesis frequency of the
lilac microcuttings of ‘Nikitskaya’ cultivar in control was from 15 to 35%, with laser
stimulation from 50 to 65%. The effect of the rhizogenesis stimulation was the most
obvious in lighting of 1,400-1,800 lux. In this case, more roots per rooted microcutting
were formed. The selection of the optimal mode and methods of supersonic irradiation
enabled the 1.5-2 increase of rooting effectiveness of the cultivars possessing lower
rooting capacity in vitro and acceleration of rhizogenesis in all studied forms.
166 166
Регулирование процесса ризогенеза при клональном
микроразмножении садовых культур
167
167
A study of the ontogenesis of early-maturing walnut cultivars in
Voronezh
Recently, much attention has been paid to early-maturing or neotenic forms of walnut
(Juglans regia L.), which start to bear nuts at the age of 2-3 years, while for ordinary forms
it takes 8-12 years. The seed reproduction of walnut has a number of advantages, since
plants adapt to the given conditions from the beginning and later they are more stable,
tolerant and drought-resistant. The research deals with ontogenesis of early-maturing
walnut and is based on ‘Ideal’ cultivar (the breeder — E.V. Evangelista). In the research, we
studied fruit characteristics: the average size of the nut was 38.6 mm × 34.0 mm, the fruit
body had a slightly ridged surface, the shell was 0.9 mm thick. 100 nuts weighed 1.0-1.2 kg.
After snow stratification, 200 nuts were sown on the trial plot in the forest nursery of All-
Russian Research Institute of Forest Genetics, Breeding and Biotechnology, Voronezh.
When monitoring the seedlings, we identified the following ontogenetic stages of walnut:
latent period (seeds); pre-generative period (seed germination and seedling growth) and
generative (fruiting phase). Shoots appeared after 10-12 days. Radicles of the embryo
formed tapering main roots. Vegetative growth ended in the middle of June with the
formation of buds. Fruiting shoots had two growth cycles: the spring one ended in June
and the summer one – in August. The generative phase of ontogenesis began in May next
year, when the average height of the plant was 14-16 cm and 40% of them formed flowers.
In September that year, the first nuts were produced. This indicates that the
acclimatization process described was successful and the cultivar can be recommended for
the establishment of early-maturing walnut plantations in the Central Black Earth Region.
168 168
Изучение онтогенеза скороплодных форм ореха грецкого в
условиях Воронежа
169
169
Dynamics of phenolic compounds in representatives of genus
Lavandula L. during the growing season
Anfisa Paliy, Valeriy Rabotyagov, Ivan Paliy, Yurii Khokhlov, Olga Startseva
Federal State Funded Institution of Science "The Labor Red Banner Order Nikita Botanical
Gardens – National Scientific Center of the RAS", Yalta, 298648, Russian Federation;
E-mail: runastep@mail.ru
The change in the content of phenolic acids and flavonoids in the leaves of Lavandula
angustifolia Mill., L. latifolia Medic. and their allotriploids L. × intermedia Emeric ex Loisel.
during the spring-summer vegetation has been studied. Total phenolics were determined
using Uv-Vis spectrophotometry. Phenolic acids and flavonoids were identified and
quantified by the HPLC method with diode-matrix detection. The content of phenolic
compounds in the leaves of L. latifolia (29.3-90.2 mg·g-1 d.w.) was found to be higher than
in L. angustifolia (27.2-56.3 mg·g-1 d.w.) and L. × intermedia (33.2-49.2 mg·g-1 d.w.).
Maximum content of total phenolics has been found during the period of mass flowering,
and minimum has been found during the period of complete regrowth of shoots. Three
phenolic acids have been identified in the leaves: neochlorogenic, trans-caffeic and
rosmarinic acid. The extracts also contain apigenin-7-O-glucoside, luteolin-7-O-glucoside
and coumarin. The maximum content of rosmarinic acid (4.6 mg·g-1 d.w.) and luteolin-7-O-
glucoside (0.1 mg·g-1 d.w.) was found in the leaves of L. latifolia at the end of the growing
season. The maximum content of neochlorogenic acid (0.3 mg·g-1 d.w.) and apigenin-7-O-
glucoside (1.7 mg·g-1 d.w.) in leaves of L. angustifolia was found at the beginning of the
growing season. In L. angustifolia and L. × intermedia trans-caffeic acid was not found. In
the leaves of L. latifolia, its content reached a maximum (1.7 mg·g-1 d.w.) at the end of the
growing season. Thus, the allotriploid L. × intermedia, in terms of the content of the sum of
phenolic compounds and individual components, mediates the original parental forms.
Obtained results show that the leaves of L. angustifolia, L. latifolia and their hybrids are
valuable sources of phenolic acids and flavonoids.
This study was funded by the research grant No. 14-50-00079 of the Russian Science
Foundation.
170 170
Динамика фенольных соединений у представителей рода
Lavandula L. в течение вегетации
Палий А.Е., Работягов В.Д., Падий И.Н., Хохлов Ю.С., Старцева О.В.
Федеральное государственное бюджетное учреждение науки «Ордена Трудового
Красного Знамени Никитский ботанический сад – Национальный научный центр
РАН»,пгт Никита, г. Ялта, 298648, Российская Федерация; e-mail: runastep@mail.ru
171
171
Realization of grape morphogenetic capacity using seeds cultured in
vitro
172 172
Реализация морфогенетического потенциала винограда в культуре
семян in vitro
173
173
Component composition of essential oil of Thuja plicata Don ex. D. Don
on the Southern Coast of the Crimea and the Black Sea Coast of the
Caucasus
Yurii Plugatar1, Oksana Shevchuk1, Vitali Leiba2, Lev Dmitriev3, Sergei Feskov1,
Vladimir Gerasimchuk1, Valeria Dmitrieva
1 Federal State Funded Institution of Science "The Labor Red Banner Order Nikita
Botanical Gardens – National Scientific Center of the RAS", Yalta, 298648, Russian
Federation; E-mail: oksana_shevchuk1970@mail.ru
2Abkhazian Research Forest Experimental Station, Ochamchira, The Republic of Abkhazia
3Russian Timiryazev State University, Timiryazevskaya Str., 49, Moscow, 127550, Russian
Federation
This study was funded by research grant No. 14-50-00079 of the Russian Science Foundation.
Keywords: Thuja plicata Donn ex D.Don, essential oil, α - and β - thujone, Southern Coast
of the Crimea, Black Sea Coast of the Caucasus
174 174
Компонентный состав эфирного масла Thuja plicata Donn ex D.Don
на Южном берегу Крыма и Черноморском побережье Кавказа
Ю.В. Плугатарь1, О.М. Шевчук1, В.Д. Лейба2, Л.Б. Дмитриев3, С.А. Феськов1,
В.Н. Герасимчук1, В.Л. Дмитриева
1ФГБУН «Ордена Трудового Красного Знамени Никитский ботанический сад –
Национальный научный центр РАН», Никитский спуск 52, пгт Никита, г. Ялта,
298648, Российская Федерация; e-mail: oksana_shevchuk1970@mail.ru;
2Абхазская научно-исследовательская лесная опытная станция, г. Очамчыра,
Республика Абхазия;
3РГАУ - МСХА им. К.А. Тимирязева, ул. Тимирязевская, 49, г. Москва, 127550,
Российская Федерация
Ключевые слова: Thuja plicata, эфирное масло, α - и β - туйон, Южный берег Крыма,
Черноморское побережье Каваказа.
175
175
Preliminary study on pomological characteristics of different mulberry
accessions in Istria, Croatia: towards their conservation
E-mail: skink@pu.t-com.hr;
2Skink Ltd, Valata bb, 52210 Rovinj, Croatia; E-mail: iva.prgomet@gmail.com;
3University of Turin, Dep. Agricultural, Forest and Food Sciences, Largo Braccini 2, 10095
Mulberry is a forgotten and underutilized fruit species which fruits are of very high
nutritional value. In Croatia, the ones that dominate are white (Morus alba) and black
(Morus nigra) mulberries. There are no planted orchards, but thousands of individual trees
are growing in private gardens or tree alleys. The aim of this preliminary study was to
register the best quality mulberry accessions in the Istria County based on their
pomological characteristics. The focus was on size and color of the fruit and resistance to
diseases and pests. Additionally, the phenophase development of selected trees was
observed. Five white (BD1, BD2, BD3, BD4 and BD5) and seven black (CD1, CD2, CD3, CD4,
CD5, CD6 and CD7) accessions were selected. Based on the measurements performed, it
was found that the average fruit weight varied from 1.35 g for CD3 to 3.76 g for CD7 within
black accessions, and from 1.42 g for BD2 to 2.05 g for BD5 within white accessions. The
shortest pedicel was observed for accessions CD2 and BD3 (5.00 mm and 4.72 mm,
respectively) and the longest ones for CD4 and BD5 (12.98 mm and 10.72 mm,
respectively). The earliest accessions to ripe were CD5, CD6, CD7 and BD4, while all the
others were fully ripened around 7 days later. Further studies, mainly genetic and
nutritional analyses, should be performed towards to conservation of mulberry accessions
biodiversity in Croatia.
176 176
Biochemical analysis of Passiflora ligularis zygotic embryo germination
after cryopreservation
Débora Prudente1, Renato Paiva1, Débora Domiciano1, Lucas Souza1, Fernanda Nery2 and
Patrícia Paiva3
1Universidade Federal de Lavras UFLA, Departamento de Biologia, 37200-000 Lavras-
177
177
Effects of selenium nanoparticles on increasing the salt tolerance of
some annual flower crops
Anna Repetskaya, Irina Yurkova, Alexander Omelchenko, Denis Panov, Irina Kazakova
V.I. Vernadsky Crimean Federal University, 4 Vernadsky Prospekt, Simferopol, 295007,
Russian Federation; E-mail: anna.repetskaya@gmail.com
One of the important indicators of the viability of plants is resistance to salt stress. Soil
salinity not only lowers crop yields, but also negatively affects ornamental plants.
Increasing the resistance of plants to salinity has both fundamental and practical
significance. An important role in enhancing the adaptive potential of plants belongs to
selenium. Selenium has antioxidant properties and is part of the structure of one of the
main antioxidant enzymes, glutathione peroxidase. Unlike ionic forms of selenium,
nanoparticles are less toxic, biologically available and have a prolonged action. However,
the effect of selenium nanoparticles on plant resistance to salt stress has been poorly
studied. The aim of this work was to study the effect of selenium nanoparticles on the salt
tolerance of annual flower crops. Selenium nanoparticles were obtained by original
technology by the reduction of sodium selenite by cysteine and the stabilization of
nanoparticles by sodium alginate. The morphology of the surface of a thin nanocomposite
selenium film was studied by scanning atomic force microscopy (AFM). It is shown that the
surface of a film is a cluster of nanoclusters consisting of nanograins. The average size of
the nanograin is about 34 nm. The research subjects were the French marigold (Tagetes
patula L.) and elegant zinnia (Zinnia elegans Jacq.). Seeds were soaked for four hours in
water solutions of nanoselenium at a concentration of 5.0; 10.0; 20.0 and 30.0 mg L-1.
Treated seeds of T. patula were germinated in Petri dishes in saline solutions (100 mM
NaCl) for 7 days at 24°C. Z. elegans seeds were grown on a saline substrate (2.0 g kg-1
NaCl) in the vegetation vessels for 30 days at 24°C. Control served as: 1 – seeds soaked in
distilled water; 2 – seeds soaked in distilled water, grown under conditions of salt stress.
The protective effect of selenium nanoparticles on seed germination, the morphometric
indices of plants (linear dimensions of roots and the aerial part, the mass of wet and dry
matter), as well as the content of proline in the conditions of simulated chloride salinity
are investigated. It is shown that the treatment of T. patula and Z. elegans seeds with
nanoselenium increases seed germination, morphometric indices of plants and proline
content in comparison with the control samples (without nanoselenium, NaCl). The
maximum effect was obtained at the concentration of nanoselenium 10.0–20.0 mg L-1, and
at 30.0 mg L-1, all the morphometric indices were suppressed. Thus, the nanocomposite of
selenium stabilized by sodium alginate at certain concentrations can activate the
protective mechanisms of flower cultures under conditions of salt stress.
Keywords: Tagetes patula L., Zinnia elegans Jacq., nanoselenium, AFM, salt stress.
178 178
Влияние наночастиц селена на повышение солеустойчивости
некоторых однолетних цветочных культур
А.И. Репецкая, И.Н. Юркова, А.В. Омельченко, Д.А. Панов, И.С. Казакова
ФГАОУ ВО «Крымский федеральный университет имени В. И. Вернадского»,
г. Симферополь, Российская Федерация; e-mail: anna.repetskaya@gmail.com
Ключевые слова: Tagetes patula L., Zinnia elegans Jacq., наноселен, АСМ, солевой
стресс.
179
179
Adaptogenic effect of Epin-extra drug on seed germination and
growth of seedlings in Cucumis sativus L. under cadmium stress
G.V. Reshetnik, S.I. Chmelеva, A.I. Sidyakin, S.F. Kotov, N.A. Sobchuk
V.I. Vernadsky Crimean Federal University, 4 Vernadsky Prospekt, Simferopol, 295007,
Russian Federation; E-mail: reshetnikgv@gmail.com
The purpose of our research was to study influence of Epin-extra drug on catalase activity
of germinating seeds and growth of seedlings in Сucumis sativus L. cultivar ‘Fenix plus’
under the effect of cadmium acetate various concentrations. It was necessary to determine
the optimal concentration of Epin-extra drug, to specify the pre-sowing seed treatment
exposure, to ascertain the catalase activity of germinating seeds and the morphometric
indicators of the seedlings. The solutions with different concentrations of cadmium acetate
(10-2 – 10-4 М, 100 µM and 250 µM) were used. 0.05% Epin-extra drug as an optimal
concentration was determed. The exposure of pre-sowing seed treatment was 4 h. Positive
antistress influence of Epin-extra drug on the germination of С. sativus. seeds under the
effect of cadmium acetate was determined. Seed germination in solutions of cadmium
acetate of various concentrations was inhibited from the first days of the experiment. At a
salt concentration of 10-2 М, the germination of the seeds was completely inhibited. At 10-
3 М, the germination rate was increased to 65%, at 10-4М – 73%. Pre-sowing seed
treatment with Epin-extra drug increased this rate on 20%. The catalase enzyme activity
was decreased due to increase of metal concentration in the medium. Pre-sowing seed
treatment in 0.05% solution of Epin-extra drug stimulated activity of the catalase enzyme
at maximum values of Cd(CH3COO)2 by an average on 10%. Under the effect of the metal
during the vegetation, the indicators of growth of the aerial part and the root system of the
cucumber seedlings were below the test ones by an average on 40%. The exogenous
epibrassinolide was increase of the aerial part on 15%, the wet weight on 17% and the dry
weight on 29%.
As a result, the studies have shown that the pre-sowing treatment of С. sativus seeds
with the growth regulator Epin-extra induces the resistance to sublethal concentrations of
cadmium, contributing to decrease of the inhibitory effects of the growth processes.
180 180
Адаптогенное действие препарата Эпин - экстра на прорастание
семян и рост проростков Cucumis sativus L. при кадмиевом стрессе
Г.В. Решетник, С.И. Чмелёва, А.И. Сидякин, С.Ф Котов, Н.А Собчук
Крымский федеральный университет имени В.И.Вернадского
проспект Вернадского, 4, г. Симферополь, 295007, Российская Федерация;
e-mail: reshetnikgv@gmail.com
181
181
Some features of Pinus sabiniana Douglas male generative matter
development in the conditions of the Southern Coast of Crimea
Tatiana Sakhno
Federal State Funded Institution of Science "The Labor Red Banner Order Nikita Botanical
Gardens – National Scientific Center of the RAS", Nikita, Yalta, 298648, Russian Federation;
E-mail: sahno_tanya@mail.ru
Digger pine (Pinus sabiniana Douglas) is an endemic species of Californian flora. It has been
under cultivation in Crimea since 1832. Digger pine occurs in the park groves of the bottom
chord of the Southern Coast of Crimea (SCC). The development characteristics of
introduced plant reproduction processes are classified as the most informative attributes
of their successful acclimatization. The development phenology and arborescent pollen
quality for genus Pinus L. representatives are the most important characteristics of plants’
state, their biological potential and response level for environmental factors’ dynamics.
Using of standard reproductive biological and biotechnological means allowed to gather
the information about the state of P. sabiniana male generative matters while introduction
in the SCC. It has been determined that in 2017-2018 P. sabiniana pollination timescales
had essential differences, going through unit phenological phases was observed in time-
shifting of 15 days and the variation in the sum of effective temperatures above 5 °C –
48.5°C. Pollen dispersion duration varies from 16 to 19 days and depends on a temperature
regime, wind speed and depth of rainfall during the period of pollination. The pollen seeds’
morphological analysis has shown that in 2018, the quantity of development anomalies
was 3.1 times larger than in 2017. Pollen vitality in 2018 has also been decreased by
21.5%. A head start of vegetation in the conditions of the SCC adversely effects on
P. sabiniana male generative matters. Huge daily average temperature swings with the
difference over 8°C while micro-sporogenesis occure expansion in the number of abnormal
pollen seeds and altogether reduce the quality of a produced pollen.
This study was funded by a research grant No. 14-50-00079 of the Russian Science
Foundation.
Keywords: Pinus sabiniana, introduction, pollen, vitality, anomalies, the Southern Coast of
Crimea.
182 182
Некоторые особенности развития мужской генеративной сферы
Pinus sabiniana Douglas в условиях Южного берега Крыма
Т.М. Сахно
Федеральное государственное бюджетное учреждение науки «Ордена Трудового
Красного Знамени Никитский ботанический сад − Национальный научный центр
РАН», г. Ялта, Республика Крым, 298648, Российская Федерация;
e-mail: sahno_tanya@mail.ru
183
183
Challenges of differentiated assessment in situ and ex situ dynamics of
vital functions of woody plants of forest ecosystems
Forest communities occupy almost a third of the world's land surface. Many scientific
studies on the problems of assessment of the negative impact of anthropogenesis on the
environment are associated with the study of woody plants as an important component of
forest biocenoses. Traditional objects of research are forest plantations growing in the
zone of exposure to atmospheric emissions of industrial facilities. The formation of the
ecological profile and the allocation of the priority directions of distribution along the
wind rose allows to characterize the dispersion gradient from a specific source of
atmospheric pollutants. It is not always possible to accurately estimate the intensity and
exposure of the pollutants in situ. This problem is to some extent solved by ex situ
observations in the laboratory. The ratio and analysis of quantitative and qualitative
results of laboratory observations allows to model and predict the state and development
of forest communities in connection with the special aspects of the dynamics of
technogenic pollution. However, the disadvantage of such studies is that the experimental
observations ex situ cannot completely take into account the full range of factors acting in
situ, as well as the fact that different biochemical active substances can have a cumulative
character of influence on organisms for a long time, manifested not only in the violation of
plant life functions during the observation period, but also in the prolonged change in the
genotypic structure of natural populations. Therefore, one of the urgent challenge is the
expansion of scientific research in the field of system assessment of the state and
prospects of forest ecosystems development on the basis of integrated use of methods of
analysis in situ and ex situ dynamics of vital functions of woody plants in connection with
man-made pollution of the natural environment.
Keywords: forest ecosystems, woody plants, pollutants, vital functions, dynamics, in situ,
ex situ.
184 184
Проблемы дифференцированной оценки in situ и ex situ динамики
жизненных функций древесных растений лесных экосистем
185
185
Conservation methods of the unique Czech gene pool of whitebeams (the
genus Sorbus)
Jana Šedivá, Roman Businský, Markéta Pospíšková, Jiří Velebil, Hana Drahošová,
Vladimír Zíka
Silva Tarouca Research Institute for Landscape and Ornamental Gardening (VÚKOZ),
Květnové nám. 391, Průhonice, Czech Republic; E-mail: sediva@vukoz.cz
The whitebeam (Sorbus sp.) is the only representative of endemic trees occurring in the
Czech Republic. At present, 14 endemic species of Sorbus are described in the Czech
Republic. Most of them are located on sites representing residual fragments of natural
habitats in the cultural landscape. A study was focused on four apomict species: Sorbus
gemella, S. omissa, S. abscondita, S. kitaibeliana, discovered recently. The aim of the project
is to develop a set of procedures for the protection of selected endemic whitebeams in their
unique natural habitats in the Czech Republic. The subject of the study is: a) assessment of
the current status of existing localities (number of individuals, habitat and vegetation
conditions, degree of disturbance and threats); b) evaluation of intrapopulation and
interpopulation genetic variability and estimation of the parentage using molecular
markers; c) determination of conditions of the in vitro propagation (composition of the
medium, conditions of cultivation and acclimatization) and generative propagation (seed
treatment, composition of substrates and germination conditions); d) extending the
existing Sorbus collection in the Dendrological Garden of VÚKOZ. The project is the first
year of the solution.
This work was supported by the Technology Agency of the Czech Republic (no. TH03030037).
186 186
Some biological features of lavender and lavandin in relation with virus
infection
Lavandula angustifolia L. is one of the valuable essential oil and ornamental plants. This is
an evergreen shrub in the height of 60-80 cm with highly branched lower vegetative
shoots and numerous young flowering shoots. In the Crimea lavender blossoms in June-
August, flowers are full, bisexual, zygomorphic, collected in false whorls, forming spicate
inflorescences. Corolla is not clearly bilabiate, up to 1 cm long, with large lobes, bluish-
violet, pubescent. Calyx is tubular, 0.4-0.7 cm long, with prongs, densely pubescent, not
falling. Pollen in lavender is 3-celled, 3-furrowed. In the total mass of pollen, L. angustifolia,
lavandin ('Rabat' and 'Snezhny Bars') and studied cultivars of lavender ('Stepnaja',
'Belyanka' and 'Rannyaya'), the pollen grains can be of various sizes, full and sterile. When
the ploidy of plants changes, the size of the pollen grains and the number of pores change.
The content of morphologically normal pollen grains in the studied taxa ranges from 30 to
90%. The highest proportion of them, exceeding 90%, was noted in L. angustifolia and
lavender of the 'Rannyaya' cultivar. The affection of the plants of lavender and lavandin by
the virus Tobacco mosaic virus (TMV) led to the appearance of yellow spots on the leaves
and their deformation, as well as to violations of internal processes of development of
male generative structures. With the determining dependence of the formation of
morphologically normal pollen grains from the genotype, the virus infection of individuals
was particularly expressed in the anomalies and deficiency of the pollen grains.
The authors are grateful to the chief scientist of the FSFIS "NBG-NSC", professor, Doctor of
Biological Sciences, V.D. Rabotyagov for the provided breeding material for research.
This study was funded by a research grant No. 14-50-00079 of the Russian Science
Foundation.
187
187
Некоторые биологические особенности лаванды и лавандина в
связи с их инфицированностью вирусами
188 188
New cherry-adapted strain of Plum pox virus - PPV-CV (Cherry Volga)
E-mail: s-chirkov1@yandex.ru
2Tatar Research Institute of Agriculture, Kazan, Republic of Tatarstan, 420059, Russian
Federation
Plum pox virus (PPV, genus Potyvirus, family Potyviridae) is considered the most
detrimental viral pathogen of stone fruit crops. Nine PPV strains (D, M, EA, C, Rec, W, T, An
and CR) recognized so far. They differ in genome sequences, antigenic and epidemiological
properties, geographical distribution, pathogenicity and host range. It was thought that
only the strains C and CR were capable of infecting sour cherry (Prunus cerasus). In 2016,
atypical PPV isolates Tat-2 and Tat-4 were detected on wild sour cherry trees and root
offshoots, displaying symptoms of Sharka disease in Tatarstan (Russia). ELISA with strain-
specific monoclonal antibodies demonstrated that both isolates reacted only with the
antibody AC to strain C. According to RT-PCR, the Tat-2 and Tat-4 were recognized only
with the primers P1/PD to the strain D. The sequences of full-length genomes of Tat-2 and
Tat-4 consisting of 9792 nucleotides were determined using the Illumina high-throughput
sequencing. They were 99.4 and 99.7% identical to each other at the nucleotide and amino
acid levels, respectively. Sequence similarities with other PPV strains were 82.6 to 83.3%
for genomic RNA and 92.6 to 93.8% for polyprotein. Phylogenetic analysis showed that
Tat-2 and Tat-4 grouped with isolates of the strains C and CR, forming a separate clade
within this cluster with 100% bootstrap support. Thus, Tat-2 and Tat-4 represent a new
PPV strain for which the name CV (Cherry Volga) was proposed. Using the strain-specific
primers targeting P1 and coat protein genes, three additional strain CV isolates (Tat-36,
Tat-39, Tat-49) were found in abandoned cherry collections and own-rooted trees of the
cultivar ‘Morel Rannyaya’. The discovery of the third cherry-adapted strain of the virus
may contribute to a better understanding of the origin, genetic diversity and evolution of
PPV as well as identification of host specificity determinants.
This work was funded by the Russian Science Foundation, grant No. 14-24-00007.
Кeywords: stone fruit crops, cherry, plum pox virus, strain CV.
189
189
Новый адаптированный к вишне штамм вируса оспы сливы - PPV-
CV (Cherry Volga)
Российская Федерация
Вирус оспы (шарки) сливы (Plum pox virus, PPV, род Potyvirus, семейство Potyviridae)
является самым вредоносным для косточковых культур. Было идентифицировано 9
штаммов вируса (D, M, EA, C, Rec, W, T, An и CR), которые различаются по
последовательности генома, антигенным и эпидемиологическим свойствам,
географическому распространению, патогенности и кругу хозяев. Считалось, что
только штаммы С и CR способны заражать вишню (Prunus cerasus). В 2016 году в
Татарстане (Россия) на дикорастущих деревьях и корневой поросли вишни с
симптомами шарки обнаружены атипичные изоляты PPV, названные Tat-2 и Tat-4,
соответственно. В иммуноферментном анализе со штаммоспецифичными
моноклональными антителами оба изолята реагировали только с антителами АС к
штамму С. При определении штамма методом полимеразной цепной реакции с
обратной транскрипцией Tat-2 и Tat-4 распознавались только праймерами P1/PD к
штамму D. Полногеномные последовательности этих изолятов длиной 9792
нуклеотида были определены методом высокопроизводительного секвенирования
на платформе Illumina. На нуклеотидном и аминокислотном уровнях Tat-2 и Tat-4
идентичны между собой на 99,4 и 99,7%, соответственно. Сходство с другими
штаммами PPV составило 82,6 - 83,3% для геномной РНК и 92,6 - 93,8% для
полипротеина. При филогенетическом анализе Tat-2 и Tat-4 группировались с
изолятами штаммов C и CR, образуя при этом c вероятностью 100% отдельную
кладу в кластере изолятов, адаптированных к вишне. На основании анализа
полногеномных последовательностей можно утверждать, что изоляты Tat-2 и Tat-4
представляют собой новый штамм вируса, для которого предложено название CV
(Cherry Volga). С помощью штаммоспецифичных праймеров, таргетирующих гены
P1 и белка оболочки, в заброшенных коллекциях и на корнесобственных деревьях
вишни сорта Морель Ранняя были обнаружены 3 дополнительных изолята штамма
CV (Tat-36, Tat-39, Tat-49). Открытие третьего штамма вируса, адаптированного к
вишне, может способствовать более полному пониманию происхождения,
генетического разнообразия и эволюции PPV, а также выявлению генетических
детерминант, определяющих круг хозяев вируса.
Ключевые слова: косточковые культуры, вишня, вирус оспы сливы, штамм CV.
190 190
Variability of microsatellite loci in ‘Antonovka’ apple landraces from
VIR collection
VIR collection of apple is the largest in Russia and includes more than 3,000 accessions.
Among apple landraces from the collection, group of ‘Antonovka’ landraces, known for its
winter hardiness and disease resistance, is of special interest. To study genetic diversity of
15 ‘Antonovka’ accessions from VIR collection, eight SSR markers were used (CH02c02b,
CH02g04, CH03d01, CH03d11, CH03d08, CH04e03, CH01f03b and CH03a04). Analysis was
performed on ABI Prism 3130xl (Applied Biosystems). As a result, 56 alleles were detected
in eight studied loci of 15 ‘Antonovka’ accessions. The number of alleles per locus varied
from five (CH02c02b, CH03d01, CH03d11 and CH04e03) to ten (CH03d08 and CH03a04),
and averaged seven. The polymorphism information content (PIC) was rather high (0.639
(CH02c02b)-0.821 (CH03d08)) with an average values of 0.706. The average expected
heterozygosity (H) was 0.740. Dice genetic similarity coefficient averaged 0.402. The
maximum similarity (not lower than 0.867) was revealed for three ‘Antonovka
obyknovennaya’ (Common Antonovka) accessions (K74, K711 and K21190), having
identical alleles in five SSR loci. A high similarity rate (0.621-0.690) was also noted
between this group of samples and ‘Antonovka Ploskaya’ (K31977). ‘Antonovka
obyknovennaya’ (K74) and ‘Antonovka Monastyrskaya’ (K68) had the same set of alleles at
all loci, and can be duplicates. The lowest level of similarity (0.069 and 0.074) was
detected between ‘Antonovka Krasnaya’ (K21652), ‘Antonovka obyknovennaya’ (K74) and
‘Antonovka Privalovskaya Zimnyaya’ (K21641). For both ‘Antonovka Pozdnyaya
Uspenskaya’ (K21645) and ‘Antonovka Zimnyaya’ (K13400), specific (for 15 studied
accessions) alleles were identified in six of the eight SSR loci studied.
Thus, the SSR loci polymorphism of 15 ‘Antonovka’ accessions was unexpectedly
high and for all studied samples, except for ‘Antonovka obyknovennaya’ (K74) and
‘Antonovka Monastyrskaya’ (K68), specific combinations of alleles were revealed. The
obtained results are part of a large-scale study of genetic diversity and certification of the
VIR apple collection.
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191
Вариабельность микросателлитных локусов сортообразцов
Антоновки народной селекции в коллекции яблони ВИР
192 192
Strategy for engineering PPV resistant cis-genic stone fruit trees by
silencing of the eIF4 host initiation translation factors
of the RAS, 6 Prospect Nauki, Puschino, Moscow region, 142290, Russian Federation;
2Lomonosov Moscow State University, Moscow, Russian Federation
E-mail: mitiouchkina@rambler.ru
Plum pox or sharka disease is the most serious viral disease affecting plum, apricot and
peach. Last decade, genetic transformation has been used as an efficient method in
engineering viral resistance for many crops, including stone fruit trees. In this study, RNA
silencing approach was used to develop Plum pox virus (PPV) resistance in plants by
manipulating the expression of eIF4 host initiation translation factors involved in the
potyvirus infectious cycle in plant species. Two DNA constructs containing self-
complementary hairpin structure of peach eIF(iso)4G and eIF(iso)4E genes under RBCS
peach promoters/terminators have been developed. To produce marker-free cis-genic
events, all vectors also included the sequence of the chemical-inducible RecLBD-CodA
system for precise removal of undesirable antibiotic resistance genes used for transgenic
tissue selection. Since the production of transgenic stone fruit plants is a difficult long time-
consuming process, a series of transgenic Nicotiana tabacum plants were firstly produced
to evaluate the efficiency of T-DNA transfer for achieving the host gene knockdown when
various host factors were used. Despite the fact that transgenic plants with both constructs
were generated, a complete sequence of eIF(iso)4E self-complementary hairpin structure
was introduced more effectively (94%), than the similar hairpin sequences of the
eIF(iso)4G gene (59%). The expression of incorporated hairpin constructs was confirmed.
The effect of silencing of eIF4 host initiation translation factors on the virus resistance will
be discussed.
Acknowledgment. This study was funded by the research grant No. 14-50-00079 of the
Russian Science Foundation.
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193
Стратегия создания устойчивых к PPV цисгенных косточковых
культур путем замалчивания генов кодирующих факторы
инициации трансляции eIF4
Сидорова Т.Н 1,2, Фролова А.3, Тимербаев В.Р.,2, Митюшкина Т.Ю.1,2, Пушин А.С.1,2,
Долгов С.В.1,2
1 ФГБУН «Ордена Трудового Красного Знамени Никитский ботанический сад –
Российская Федерация;
e-mail: mitiouchkina@rambler.ru
Болезнь Шарки (PPV, Plum Pox Virus) представляет собой одно из наиболее
вредоносных вирусных заболеваний, которое поражает деревья сливы, абрикоса и
персика. В последнее десятилетие для достижения устойчивости к вирусным
заболеваниям различных видов растений, включая косточковые культуры, активно
используют методы генетической трансформации. В данном исследовании
изучается стратегия создания вирусоустойчивых цисгенных растений, в которых с
помощью метода РНК интерференции ингибируется экспрессия генов кодирующих
факторы инициации трансляции растений eIF4, использующихся вирусами
семейства Potyviridae для завершения своего инфекционного цикла. Нами созданы
векторные конструкции, которые содержат самокомплементарные шпилечные
последовательности генов, кодирующих факторы eIF(iso)4G и eIF(iso)4E персика,
под промотером и терминатором RBCS персика. Для получения цисгенных растений,
свободных от ненужных трансгенных последовательностей, векторы также
содержат последовательность химически-индуцируемой системы RecLBD-CodA,
позволяющей удалить гены устойчивости к антибиотику, используемые для отбора
растений после генетической трансформации. Поскольку получение трансгенных
растений плодовых культур является сложным и долговременным процессом, для
оценки эффективности переноса Т-ДНК и нокдауна целевых генов при
использовании созданных конструкций, нами сначала получен ряд трансгенных
растений Nicotiana tabacum. Несмотря на то, что использование обеих конструкций
позволило успешно получить трансгенные растения, перенос полноценных
шпилечных последовательностей фактора eIF(iso)4E проходил эффективнее (94%),
чем последовательности фактора eIF(iso)4G (59%). Подтверждена успешная
экспрессия шпилечных конструкций перенесенных в геном растений. Проведена
оценка РНК-интерференционного ингибирования генов факторов инициации
трансляции eIF4 растения-хозяина для изучения устойчивости к вирусу.
194 194
Content of biologically active substances in fruits of hybrid peach plants
from breeding pool of the Nikita Botanical Gardens
For several decades, Nikita Botanical Gardens has been working on breeding new
promising peach cultivars. New cultivars must exhibit better characteristics than the
existing ones. Modern hortculture needs peach cultivars with such properties as good
commercial qualities of fruits, early bearing age, resistance to diseases, high and stable
yields. High-quality fruits must have an attractive appearance, excellent taste properties,
ensure ease of transport and have a high content of biologically active substances. The
solution of these problems makes possible the obtaining of economically high efficient
cultivars for implementation into industrial horticulture. The breeding process included
several researches on detecting biologically active substances in mesocarp of peach plants
studied, which were obtained through purposeful hybridization. The analysis showed that
the solids content in fruits of researched hybrid plants varied between 13.30% (in the
seedling 7/43) to 19.70% (7/43). The fruit flesh showed a low content of ascorbic acid
(6.60 to 13.20 mg/100 g). Its maximum content emerged in fruits of the hybrid form 2/18.
The total number of organic acids in fruits varied from 0.43 % (7/73) to 0.57% (7/43).
The total concentration of phenolic compounds in fruits varied from 150 mg/100 g (4/43)
to 465 mg/100 g (7/43). Maximum concentration of leucoanthocyanins in fruits showed
the peach seedling 7/43 (456 mg/100 g) and of flavons – the seedling 4/43 (144 mg/100
g). The seedling 7/43 proved to be promising for its further investigation and involving in
the breeding, thanks to maximum complex content of biologically active substances in
fruits.
This study was funded by the research grant No. 14-50-00079 of the Russian Science
Foundation.
195
195
Содержание биологически активных веществ в плодах гибридных
растений персика селекционного фонда Никитского ботанического
сада
196 196
Biotechnological methods of maintaining collections of the genus
Ribes L.
The development of effective methods for plant clonal micropropagation is a basic and
necessary condition for creation of genebanks in vitro. Currently, the gene pool collection
of samples of cultivated plants in vivo (Federal State Budgetary Scientific Institution All-
Russian Horticultural Institute for Breeding, Agrotechnology and Nursery) genus Ribes L.
has 295 cultivars: 137 – black currants, 45 – red currants, 82 – gooseberries, 31 – golden
currants. Genetic collection in vitro of black currant includes the following cultivars:
‘Barmalej’, ‘Bryanskij’, ‘Agat’, ‘Gamayun’, ‘Kudesnik’, ‘Mif’, ‘Podarok’ ‘Veteranam’, ‘Strelec’,
‘Uslada’, ‘Charodej’ and ‘Yadrenaya’; golden currant – ‘Businka’. The conservation of in
vitro gene pool collection is cost-effective for the maintenance of biological diversity, but
requires periodic testing for the genetic identity of the cultivar.
The deposition is carried out in the conditions of the cooling chamber (6±2°C), at 6-8
h photoperiod and illuminance of 500-1,000 lux. In vitro collection of currant is annually
updating on 2-3 cultivars, free from malicious viruses: Arabis mosaic virus (ArMV),
Raspberry ringspot virus (RpRSV), Tomato black ringspot virus (TBRV), Strawberry latent
ringspot virus(SLRSV) and Raspberry bushy dwarf virus (RBDV).
At the stage of explants, introduction MS culture medium was used with growth
regulators, such as 6-benzylaminopurine (6-BAP), kinetin and zeatin in diferent
concentrations (from 0.5 to 2.0 mg L-1) by step 0.5. The optimal variant was a medium with
6-BAP in a concentration of 0.5 mg L-1 for black currant and 1.0 mg L-1 for golden currant.
The survival rate of black currant explants varied within 71.3-89.7%, golden currant –
37%. The multiplication rate among the black currant cultivars ranged from 1:2
(‘Gamayun’) to 1:4 (‘Bryanskij’, ‘Agat’), golden currant – 1:6. 6-BAP concentration increase
to 2.0 mg L-1 in culture medium increased the multiplication rate of golden currant to 1:8,
but led to the appearance 35% hyperhydrated microshoots.
197
197
Биотехнологические приемы поддержания коллекций рода Ribes L.
198 198
Creation of marker-free transgenic tomato plants expressed the
supersweet protein thaumatun II
Vadim Timerbaev 1,2,3, Anna Okuneva3, Alexander Pushin 1,2,3, Sergey Dolgov 1,2,3
1Federal State Funded Institution of Science "The Labor Red Banner Order Nikita Botanical
Gardens – National Scientific Center of the RAS", Yalta, 298648, Russian Federation
2The Branch of M.M. Shemyakin and Yu.A. Ovchinnikov Institute of Bioorganic Chemistry
of the RAS, Prospect Nauki, 6, Puschino, Moscow region, 142290, Russian Federation
3All-Russia Research Institute of Agriculture Biotechnology, Timirayzevskaya Str., 42,
Production of cisgenic and intragenic marker-free plants with new traits is a modern
approach that dictates the realities of the 21st century that answer the biosafety
requirements. In our work we used the pMF1 vector containing the recombinase R and a
CodA-nptII bifunctional selectable gene for produce tomato plants carrying the supersweet
thaumatinII gene from tropical plant under the control of tomato fruit-specific ELIP or E8
gene promoter and tomato RuBisCo terminator. After agrobacterium transformation, two
strategies were followed for the selection of marker-free transgenic tomato plants. In the
early negative selection approach, from a total of 116 plants and 83 shoots was non-
transgenic escapes, 32 contained nptII gene fragment and only one marker-free plant with
excised DNA appeared. In the alternative delayed strategy, we have obtained 170
transgenic tomato lines. Then, we chose 35 transgenic tomato lines containing full T-DNA
and recombinase activity induced in explants. About half of them did not produce
regenerants on medium with 5-FC. One hundred twenty one resistant sublines were
obtained from 18 original lines. Most of them lost resistance to kanamycin in spite of the
sequence of nptII gene detected by PCR in 120 plants. Only one fully marker-free
transgenic line was obtained. We supposed that an incomplete excision and chromosomal
rearrangements due to the presence of multiple or partial T-DNA insertions occurred in
other cases. With the Southern blot analysis, we confirmed that a total of completely
marker-free transgenic tomato plants were obtained by two different selection strategies.
The thaumatin II gene expression has been detected by RT-PCR, Western blotting and
organoleptic analyses.
This study was funded by the research grant No. 14-50-00079 of the Russian Science
Foundation.
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199
Создание безмаркерных трансгенных растений томата,
экспрессирующих суперсладкий белок тауматин II
Национальный научный центр РАН», Никитский спуск 52, пгт Никита, г. Ялта,
298648, Российская Федерация
2Филиал института биоорганической химии им. М.М. Шемякина и
200 200
Comparative evaluation of introduced nectarine cultivars with a model
cultivar
The study had been carried out in 14 consecutive years (2004-2017) at the Nikita
Botanical Gardens. As a control cultivar for comparison, model one was used. The
objectives were phenological, pomological, morphological and physiological
characteristics of 25 cultivars. The researches included parameters of introduced cultivars
in comparison with similar parameters of the model one according to 15 characteristics: a
number of days starting from the agreed date (March, 1) till the beginning of blooming,
degree of blooming, duration of blooming, winter resistance, drought resistance, tolerance
to leaf-curl mosaic and mildew, the year of the first fruiting, crop capacity, fruit weight,
flesh consistency, percentage of integumentary color and fruit attraction, flesh taste and its
ability to separate from the stone. The cluster analysis was carried out to compare 25
introduced nectarine cultivars. This method made it possible to reveal similarity between
all these cultivars and model of a cultivar based on Euclidean distance. All objects were
united into classification tree that is reproduced in dendrogram. According to similarity of
characteristics and their values the following cultivars, combined into clusters (groups),
are rather close: 1. ‘Sorok let Uzbekistana’, ‘Early Rivers’, ‘Sladkoyadernyiy’, ‘Grande’,
‘Natalie’, 2. ‘Nectared C-3’, ‘Kzil Shalili’, ‘Big Top’, ‘Crimson Gold’, ‘Nectacrest’, ‘Nectared
306’, ‘Skif’, ‘May Red’, ‘Silvery’, ‘June Red’, ‘Fantasia’, ‘Stark Sanglo’, ‘Geokchayskiy 229’, ‘Le
Grande’, ‘May Grand’, ‘Super Crimson Gold’, ‘Stark Sunglo’, ‘Nectalate’; 3. ‘Kando’ and ‘Fire’.
According to complex of characteristics, 4 cultivars were marked out having the largest
similarity with the model of cultivar: ‘Geokchayskiy 229’, ‘Le Grande’, ‘Natalie’ and
‘Nectared C-3’. At the same time interrelations of economically valuable characteristics
were identified having used paired correlation coefficients. The highest correlation
coefficient was recorded between winter and drought-resistance (r=0.7), beginning of
blooming and its degree (r=0.71), fruit weight and taste (r=0.64) and etc.
This study was funded by a research grant No. 14-50-00079 of the Russian Science
Foundation.
201
201
Сравнительная оценка интродуцированных сортов нектарина с
моделью сорта
202 202
The antioxidant activity of leaves in Prunus avium at the growing clonal
stocks
The stock determines the peculiarities of the top system provision with water and
nutritional chemicals causing changes in the plants growing intensity and photosynthesis,
in the metabolic process activity. The timeliness of stock influence on the grafted plants
biochemical status increases because of the necessity of hybrid stocks and cultivars
compatibility early detection. The purpose of the research was to study the total
antioxidant activity (TAA) at new sweet cherry breeds while grafting on clonal stocks. The
research objects are three sweet cherry cultivars (‘Fatezh’, ‘Tyutchevka’ and
‘Chermashnaya’), grafted on eight clonal stocks (‘Kolt’, ‘Moskoviya’, ‘Izmaylovskiy’, ‘Stepnoy
rodnik’, ‘AVCH-2’, ‘VSL-2’, ‘VTS-13’, ‘V-5-88’). The total antioxidant activity of methanol and
liquid extracts was determined on the spectrophotometer Helios Y via DPPH method as the
extinction correlation at reaction behavior within ten min. Antioxidant activity depended
on the cultivar, stock and extracted substance. While grafting on ‘Moskoviya’ and
‘Izmaylovskiy’ stocks all studied cultivars had maximum values of antioxidant activity. The
strong positive correlation was determined between TAA values of ‘Izmaylovskiy’ stock
and grafted on it sweet cherry plants (r=0.95). Consistently high values of TAA are
examined at scion-stock combinations of ‘Fatezh’ cultivar (at average 74.,9%) and the most
winter resistant ‘Moskoviya’, ‘Izmaylovskiy’, ‘VTS-13’, ‘AVCH-2’ and ‘Stepnoy rodnik’ stocks
(78-88%). Consistently low values of TAA were at sweet cherry plants grafted on ‘V-5-88’,
‘Kolt’ and ‘VSL-2’ stocks (55-65%) It may be connected with more low adaptability of
stocks and grafted on them plants. On ‘AVCH-2’ and ‘Stepnoy rodnik’ stocks the radical
decrease of TAA at ‘Tyutchevka’ and ‘Chermashnaya’ cultivars and TAA high value at
‘Fatezh’ cultivar were marked, that showed the different compatibility of stocks with
cultivars. Antioxidant activity in sweet cherry leaves varied greatly depending on the
cultivars and stocks.
203
203
Антиоксидантная активность листьев Prunus avium при
выращивании на клоновых подвоях
204 204
Efficiency of introduced sweet cherry cultivars used in breeding
The genofund collection of sweet cherries (Prunus avium L.)in the Nikita Botanical Gardens
includes about 400 samples from 19 countries. According to their ecological and
geographical origin, the cherry cultivars belong to different groups: West European
(14.8%), American (8.5%), Caucasian (1.8%), Moldavian (1.8%), Crimean (27.4% %),
Central Ukrainian (25.6%), northern (17.5%) and Central Asian (0.4%). Cultivars vary
greatly in morphological, biological and economically valuable characteristics. A significant
range of variations in these features was determined. Genotypes - sources of economically
valuable features were involved in breeding in order to create new valuable cultivars. A
hybrid fund of more than 500 samples from 12 hybrid families was studied during 1999-
2017. The aim of our investigations was to determine the effectiveness of the introduced
cultivars use in sweet cherries breeding and selection. Twenty-nine promising hybrid
forms were selected for improving the sweet cherry assortment. Cultivars of early and very
early ripening terms were obtained using the method of embryo culture in vitro: ‘Uslada’,
‘Vesnyany Naspyvy’, ‘Prizerka’, etc. The greatest numbers of promising forms were
obtained with the participation of introduced cultivars, such as ‘Bigarreau Starking’,
‘Bigarro from Vinoli 2’ and ‘Drogana Zheltaya’. These cultivars were the best to transmit
the features of high yield, large size and good taste of fruits. The most effectiveness was a
family which the ‘Bigarreau Starking’ cultivar was used as the maternal form, and the
cultivar ‘Krupnoplodnaya’ was used as the paternal form. They produced the greatest
number of promising forms with large high-quality fruits with various terms of ripening
and high yield. Among these forms we selected the cultivars ‘Karadag’ and ‘Vytivnytsa’ that
have already been included in the Register of Plants which were suitable for distribution.
The promising cultivars, ‘Dolgozhdannaya’ and ‘Triumf’ have been transferred to the
Commission for Selection Achievements Test.
This study was funded by the research grant Nо. 14-50-00079 of the Russian Science
Foundation.
Keywords: Prunus avium L., genofund collection, introduced cultivars, breeding, hybrid,
promising form, cultivar.
205
205
Эффективность использования в селекции интродуцированных
сортов черешни
206 206
Investigation of antimicrobial activity of pea (P. sativum L.) NCR
peptides towards plant endophytic bacteria Bacillus sp. and Rahnella sp.
Ekaterina Vasileva, Marina Kliukova, Alexey Afonin, Gulnar Akhtemova, Vladimir Zhukov,
Igor Tikhonovich
Federal State Budget Scientific Institution All-Russia Research Institute for Agricultural
Microbiology (ARRIAM), Podbelsky ch., 3, Pushkin, Saint Petersburg, 196608, Russian
Federation; E-mail: evasilieva@arriam.ru
Legumes are capable of forming arbuscular mycorrhiza with Glomeromycota fungi and
nitrogen-fixing symbiosis with nodule bacteria (rhizobia). Endophytes were found in other
internal tissues of legumes, causing asymptomatic infection or having a positive effect on
growth and development of a host-plant. This types of interactions are controlled by plant
antimicrobial proteins – defensins. In some legumes defensin-like nodule-specific cysteine-
rich peptides (NCRs) were found. NCRs differently affects bacteria that penetrate into the
nodule: under the action of NCR-peptides "compatible" nodule bacteria irreversibly
differentiate into symbiotic forms – bacteroids, and "incompatible" bacteria get eliminated.
In the genome of the model legume Medicago truncatula about 700 NCR-peptides were
found, while only 8 were described in garden pea (Pisum sativum L.). Through the analysis
of metatranscriptomic data of P. sativum, we identified 715 sequences related to NCR-
peptides. Such variety of NCRs can exhibit antimicrobial properties towards bacteria
potentially capable of penetration into the nodule (endophytic bacteria). The aim of this
study is to search for and test NCR-peptides of P. sativum, demonstrating antimicrobial
activity against endophytic bacteria Rahnella sp. and Bacillus sp. To select NCR-peptides
having antimicrobial activity, various physicochemical parameters were analyzed: charge,
hydrophobicity, the Boman index and the expression level of NCR-peptide genes in the
wild-type pea line SGE. Using the algorithms of the CAMP resource (Collection of
antimicrobial peptides), the probability of antimicrobial activity was predicted. Based on
this data, candidates for chemical synthesis were selected. To confirm the predicted
antimicrobial activity, members of the endospheric community were selected (Rahnella sp.
and Bacillus sp.), which demonstrated the growth-promoting ability in a test on watercress
salad (Lepidium sativum L.).
The results of microbiological experiments to determine the elimination, minimum
inhibitory and minimum bactericidal concentration of synthesized NCR-peptides have
been shown.
207
207
Исследование антимикробной активности NCR-пептидов гороха
посевного (P. sativum L.) против растительных эндофитных
бактерий р. Bacillus и Rahnella
Е.Н. Васильева, М.С. Клюкова, А.М. Афонин, Г.А. Ахтемова, В.А. Жуков,
И.А Тихонович
ФГБНУ ВНИИ Сельскохозяйственной Микробиологии, шоссе Подбельского, 3,
196608, Пушкин, г. Санкт-Петербург, Российская Федерация;
e-mail: evasilieva@arriam.ru
208 208
Differences in peroxidase isozyme spectrums of diverse rose garden
group regenerants
The genetic similarity between regenerants and their original plants from which primary
explants were obtained is crucial due to the possible occurrence of somaclonal variation
influenced by in vitro conditions. Genetic similarity is also important as a result of unclear
membership of rose diversity in a specific garden group. Analysis of some rose groups was
conducted based on peroxidase (PO) isozyme spectra. PO is an enzyme which can be used
to identify differences between one plant species from the other, as well as the stage of
plant regeneration and their ontogenetic age. Explants of Rosa canina L., climbing cultivar
‘Aloha' and somacloned cultivar ‘Gloria Dei’ with lost ability to bloom were used as a
control. These two PO activity zones are characteristic of that part of gel with R f 0.36-0.40
which are observed in all analyzed groups: tea-hybrid forms (8 cultivar), grandiflora (2
cultivar), floribunda (2 cultivar). The dogrose PO spectra differ significantly (upward)
from the spectra of tea-hybrid and grandiflora group regenerants. Differences in isozyme
spectra depend on the regenerants' leaf location. Thus, in the ‘Komsomolskij Ogonjek’ the
upper leaf spectrum consists of 4 bands and the lower leaf reveals 5 bands. Differences are
also observed within the same group: the cultivars ‘Dame de Coeur’ and ‘Nicole’ reveal 2
zones of activity unlike cultivar ‘Folklore’ (5 zones) of the same garden group. Callus
tissues of different roses reveal an expanded activity spectrum (8-10 zones) decreasing
along side with tissue differentiation. This phenomenon is determined by the changes in
protein and hormone cell content. Adapted to non-sterile conditions, regenerants either
completely lose their isozyme spectrum (cultivar ‘Nicole’) or detect significantly reduced
spectrum in the region (cultivar ‘Folklore’). Domestic roses have wider spectra compared
to introduced roses. The differences in spectra are probably due to the evolutionary origin
of roses. Updated roses reveal a reduced spectrum.
209
209
Различия в изоферментных спектрах пероксидазы в регенерантах
садовых групп роз
210 210
Anti-stress properties of strongly diluted solution of 2-carboxy-2-
(N-acetylamino)-3-(3,5'-di-tert-4-hydroxephenyl)-potassium
propionate
The main sources of reactive oxygen species (ROS) under stress conditions are
mitochondria and chloroplasts. It has been found that the damaging effect of ROS on
respiratory activity is associated with the peroxidation of membrane lipids and, first of all,
cardiolipin and the oxidation of thiol groups of proteins, which leads to swelling of the
mitochondria, the release of cytochrome C, and possibly to the induction of apoptosis. It
could be assumed, that the preparations having adaptogenic properties should primarily
influence the generation of ROS. Antioxidants are primarily claimed to this role. The aim of our
work was to study the effect of various concentrations of potassium anphen (2-carboxy-2-(N-
acetylamino)-3-(3',5’-di-tert-butyl-4'-hydroxyphenyl), potassium propionate) on the
functional state of the mitochondria of 5-day etiolated pea seedlings (Pisum sativum L., cultivar
‘Alpha’) and computer modeling of the structure and properties of the molecular ensembles of
potassium anphen with the participation of water molecules. To study the anti-stress
properties of the preparation, the mitochondria were incubated in a hypotonic environment,
which led to a 3-4 fold increase of the fluorescence intensity of lipid peroxidation (LPO)
products in mitochondrial membranes. The drug in concentrations 10-6-10-8, 10-13 and 10-15 M
decreased the fluorescence intensity of LPO products to the control level. At the same time, it
increased the maximum oxidation rates of nicotinamide adenine dinucleotide (NAD)-
dependent substrates by 35% (10-6 M) and 44% (10-13 M). Since the mitochondria of
germinating seeds are characterized by a low rate of oxidation of NAD-dependent
substrates, the increase of the activity of NAD-dependent dehydrogenases helps to
maintain a high level of energy metabolism in the cell, which may provide the plant
resistance to stress factors. The dose dependence of the biological activity of the
preparation can apparently be explained by the possibility of the existence of kinetically
independent hydrates containing agglomerates (structures) in an aqueous medium. The
reactivity of such agglomerates is associated with the change in entropy and free energy,
depending on the structure of the solvation shell of the ligand.
211
211
Антистрессовые свойства сильно разбавленных растворов 2-
карбокси-2-(n-ацетиламино)-3-(3’,5’-ди-трет-бутил-4’-
гидроксифенил)-пропионата калия
212 212
Identification of sequences coding NCR peptides in de novo assembled
nodule "metatranscriptome" of Pisum sativum L.
Evgeny Zorin, Marina Kliukova, Alexey Afonin, Vladimir Zhukov, Igor Tikhonovich
Federal State Budget Scientific Institution All-Russia Research Institute for Agricultural
Microbiology, Podbelsky ch., 3, Pushkin, Pushkin, Saint Petersburg, 196608, Russian
Federation; E-mail: kjokkjok8@gmail.com
213
213
Идентификация последовательностей NCR пептидов в собранном
de novo “метатранскриптоме” клубеньков Pisum sativum L.
Е.А. Зорин, М.С. Клюкова, А.М. Афонин, В.А. Жуков, И.А. Тихонович
Федеральное государственное бюджетное научное учреждение “Всероссийский
научно-исследовательский институт сельскохозяйственной микробиологии”, шоссе
Подбельского, д. 3, Пушкин, г. Санкт-Петербург, 196608, Российская Федерация;
e-mail: kjokkjok8@gmail.com
214 214
АЛФАВИТНЫЙ УКАЗАТЕЛЬ АВТОРОВ
222 222
Для заметок
Научное издание
Материалы
VIII Международной научно-практической конференции
«Биотехнология как инструмент сохранения
биоразнообразия растительного мира
(физиолого-биохимические, эмбриологические,
генетические и правовые аспекты)»
В авторской редакции