COST Technical Committee "Agriculture, Biotechnology and Food

Science"
Proposal for a new COST Action

Cryopreservation of crop species in

Europe
ACRONYM: CRYOPLANET
Proposing country: Belgium
Action Proposer : Bart Panis

Plant Genetic Resources: Current situation

Europe
64 endemic plants of Europe have become extinct in recent decades
24% of the species/subspecies of certain groups of European plants
are in danger to be lost
Agricultural intensification has reduced the area under wetlands in
Europe by some 60 % in the last decades with consequent
threats on biodiversity

Worldwide

100,000 plants representing 1/3rd of plant species are

threatened
Since the 1970s large number of land races and wild
relatives are sampled and stored ex situ
Now, about 6 million samples are held in national, regional,
international and private genebank collections

Storage of Plant germplasm

In situ : Conservation in normal habitat
rain forests, gardens, farms

Ex Situ :
Field collection, Botanical gardens
Seed collections
In vitro collection
Normal growth
Slow growth (temp, O2 , H2O , medium ~)
Cryopreservation (-196C)

(DNA Banks)

What is cryopreservation ?
What ?
Storage of living tissues at ultra-low temperatures (-196C)
Use
Conservation of plant germplasm
Vegetatively propagated species (root and tubers, ornamental, fruit
trees)
Recalcitrant seed species (Howea, coconut, coffee)

Conservation of tissue with specific characteristics

Medicinal and alcohol producing cell lines
Genetically transformed tissues
Transformation/Mutagenesis competent tissues (ECSs)

Eradication of viruses (Banana, Plum)

Conservation of plant pathogens (fungi, nematodes)

What is the problem with plant

cryopreservation ?

Cryopreservation procedures are now available for about 150200 different plant species
But for each species and tissue type, the cryopreservation
protocol needs to be empirically adapted in function of their
natural freezing resistance
explant size and type
water content
Most of the work on cryopreservation of plants has been
performed in the framework of academic studies and involves
only one or a few genotypes. Only few plant germplasm
collections stored in liquid nitrogen currently exist (with a relatively
limited amount of accessions).

No real standardised methods are available !!!

Examples of existing collections in liquid

nitrogen?

National Seed Storage Laboratory (NSSL) (Fort Collins,

Colorado, USA): 2,100 accessions of apple (dormant buds)
National Clonal Germplasm Repository (NCGR) of Corvallis
(USA): 104 accessions of pear (shoot tips);
International Potato Centre (CIP) (Lima, Peru) : 345 potato
accessions
Tissue Culture BC Research Inc.(Vancouver, BC, Canada) : 5000
accessions representing 14 conifer species
AFOCEL (Association Fort Cellulose) of France, with over 100
accessions of elm (dormant buds);
National Institute of Agrobiological Resources (NIAR) of
Japan, with about 50 accessions of mulberry.
IRD (Montpellier, France) : 80 accessions of oil palm
German Collection of Micro-organisms and Cell Cultures
(DSMZ) (Braunschweig, Germany) : 519 old potato varieties
INIBAP, Laboratory of Tropical Crop Improvement,
K.U.Leuven (Heverlee, Belgium) : 440 banana accessions

Why is cryopreservation not more widely

applied?
(i) the unavailability of efficient and robust cryopreservation
protocols applicable to many plant species and diverse
germplasm types
(ii) limited awareness of plant researchers unacquainted to
recent developments in cryogenic storage methods
(iii) lack of coordinated research on plant cryopreservation.
COST action like CRYOPLANET could make the difference

Why are the objectives of CRYOPLANET ?

Objective 1: To screen in detail the current utilization of

plant cryopreservation in Europe.
Objective 2: To screen and compare the efficiency of
existing plant cryopreservation protocols.
Objective 3: To improve fundamental knowledge about
cryoprotection through the determination of physicobiochemical changes associated with tolerance
towards cryopreservation.
Objective 4: To develop new plant cryopreservation
protocols.
Objective 5: To assure the genetic stability and true-totypeness of plants after cryopreservation.
Objective 6: To apply cryopreservation to European
plant germplasm collections.
Objective 7: To proof the environmental, social and
economic impact of plant cryopreservation.

Scientific programme of CRYOPLANET ?

WG1
Fundamental aspects of
cryopreservation/cryoprotectio
n and genetic stability
Optimisation

Feed back

WG2
Technology, application and
validation of plant
cryopreservation

WG1: Fundamental aspects of

cryopreservation/cryoprotection and genetic stability
1.1. Fundamental aspect of cryopreservation and
cryoprotection
Elucidation of the physico-biochemical background of
cryoprotection and cryopreservation.
Water thermal
behavior

Proteins

Sugars

Membrane
components

Polyamines

Cytoskeletal
protein

Oxidative
stress

WG1: Fundamental aspects of

cryopreservation/cryoprotection and genetic stability
1.2. Genetic stability and authenticity
Assessment of the genetic integrity of plants to determine if
they are true to type after cryopreservation.

Assessments of phenotypic variation (morphological descriptors)

Cytological techniques to detect various types of chromosomal
instability. (polyploidy, aneuploidy and other mitotic abnormalities)
Biochemical metabolite/protein (isozyme) profiles
Genomic DNA sequences that can be analysed using a range of
hybridization and (PCR) techniques
Epi-genetic variation in chromatin and DNA methylation of gene
sequences

WG2: Technology, application and validation of plant

cryopreservation
2.1. Technology aspects of cryopreservation
Applications of different cryopreservation protocols to different
plant species and tissues.

WG2: Technology, application and validation of plant

cryopreservation
2.2. Impact and applications of cryopreservation in
plants
genebanks, establishment of cryo-bank and dissemination of
results

Organisation of CRYOPLANET
The Management Committee (MC)

Appointment of Action Chair, Vice-Chair(s) and WG Coordinators.

Planning of MC meetings and of Scientific Meetings and
Workshops.
Assessment and report of the progress made by the
different WGs
Promotion of co-operation and of data exchange between
the WGs.
Promotion and approval of Short-Term Scientific Missions,
Preparation of the Annual Reports.
Establishment and update of a Web site
Organization of contacts and common workshops
Preparation of a EU 7th framework project

Organisation of CRYOPLANET
Two Working Groups
WG1 : Fundamental aspects of
cryopreservation/cryoprotection and genetic stability
WG2: Technology, application and validation of plant
cryopreservation
Working Group meetings will be organised on a yearly base
WG coordinators
Planning the appropriate Scientific Meetings.
Coordination of the activities within the WG.
Promoting joint research (f.e. STSMs) and common
publications.
Report on the WG progress to the Action Chair and MC
Participation in the plenary and restricted MC meetings.
Inter-COST Workshops
Short-Term Scientific Missions (STSM),

Time table
Year 1

Year 2

Year 3

Year 4

Coordination
Kick-off
meeting
Homepage
Reporting
MC meeting
WG1 meeting
WG2 meeting
Workshop*
STSMs
Final Conference

MC meeting: Management committee meeting; WG meeting: Working group

meeting; STSMs: Short-term scientific missions; Workshop*: Timing of the InterCOST Workshops will be defined in agreement with the Management committee of
that specific Action.

Economic dimensions
17 COST countries
64 researchers
50 research
institutes

Dissemination

Articles in refereed scientific journals

Common reviews, books
A public website (information about the project, the
achievements, services/consultancies offered and
announcements of training workshops.
Information on the official webpages of the
collaborating institutions.
The consortium will organize workshops for scientists,
germplasm curators regulatory bodies and policy makers
At the end of the Action, the consortium will offer its
expertise as a service to the EU.
Presentations at International Conferences, for
promoting the European know-how and increasing the
international collaboration.
Teaching activities in Universities at undergraduate and
post-graduate level. Young scientists and engineers will thus
be trained and informed on the latest developments in
cryopreservation.

Acknowledgements
Carpentier Sebastien, Druart Philippe, Geelen Danny, Swennen
Rony, Tsvetkov Ivaylo, Bilavcik Alois, Faltus Milos, Zamecnik Jiri,
Antonius-Klemola Kristiina, Hggman Hely, Nukari Anna, Rokka
Veli-Matti, Uosukainen Marjatta, Dussert Stphane,
Engelmann Florent, Grapin Agns, Harvengt Luc, Malaurie
Bernard, Pros Jean-Pierre, Trontin Jean-Franois, Hfer Monika,
Keller Joachim, Meier-Dinkel Andreas, Pinker Ina, Schumacher
Heinz Martin,Benelli Carla, Caboni Emilia, Damiano Carmine,
Dulloo Ehsan, Lambardi Maurizio, Hausman Jean-Franois, Criel
Bram, Harrouni Cherif, de Klerk Geert-Jan, Raemakers Krit, Bach
Anna, Lisek Anna, Mikua Anna, Niedzielski Maciej, Pukacki
Pawel M., Rybczyski Jan J., Sochacki Dariusz, Zimny Janusz,
Miguel Clia, Reis Moura Isabel, OliveiraMargarida,Halmagyi
Adela, Palada Magdalena, Rakosy-Tican Elena, Gavrilenko
Tatjana, Salaj Terezia, Gonzlez-Benito Elena, Revilla M. ngeles,
Vieitez Ana, Pques Marc, Feki Lotfi, Aylin Ozudogru Elif, OzdenTokatli Yelda, Benson Erica, Cripps Ryan, Grout Brian, Harding
Keith, Lynch Paul T., Pritchard Hugh W.,Wetten Andy