Expression and Evolution Analysis of BCCP Family from Glycine Max and CHI Family from Arachis

Hypoga
Author :LiuYu
Tutor:WangXingJun
School :Shandong Normal University
CLC :S565.1
TYPE :Masters thesis
Download the PDF Full Text:http://www.topresearch.org/showinfo-178-678026-0.html
Year:2014
Abstract:
The existence of gene family has great significance to plant evolution. Multiplecopies of gene could
provide more adaptation opportunities and lower purifyingselection pressure. The appearance of gene
family could affect the function of organsand plant traits by affecting the expression level of proteincoding genes. Whatcontribution the gene family has done to protein translation and how the temperal
andspatial expression of the gene family is regulated are interesting. In regulation of genefamily
expression, whats the contribution from the promoters of these genes? In thisstudy, we analyzed the
expression of soybean BCCP gene family by promoter andqRT-PCR. The transcripts abundance,
promoter regulation element and activaty,methylation and evolution of this gene family were
performed.BCCP is a subunit of acetyl coenzyme A carboxylase, which is the keyrate-limiting enzyme
of fatty acid synthesis. One purpose of this study is toinvestigate expression patterns of different
member of BCCP gene family during seeddevelopment. Through the soybean genome database,
Promoter sequence informationof seven members of soybean BCCP gene family was obtained. All
these promoterswere cloned from soybean by PCR. Promoter-GUS expression vectors
wereconstructed and were transffered into tobacco plant through Agrobacterial mediatedmethod.
Promoter activaty in different tissues of transgenic tobacco was analyzed byGUS staining. Promoter
p19g03530showed strong and constitutive transcriptionalactivity in transgenic plants, and GUS
activity was observed in seeds, and plantdevelopmental stages. Promoter p18g47850showed high
activities in seedtorpedo-shape embryo period relatively. In transgenic tobacco plants,
promoterp18g47850showed
low
activities
in
seed
torpedo-shape
embryo
period.
Promoterp13g44040showed low activities in cotyledon at torpedo-shape embryo satge. Mature leavies
from transgenic plants were taken for GUS staining, and the result showed thatpromoters of BCCP
family genes has different activities in leaves. Other transgenicplants are still growing. Real-time
quantitative PCR was carried out to examine theexpression of all these soybean BCCP genes. The
results showed that the expressionlevel of BCCP19g03530are distinct high in seeds than in other
tissues.Chalcone-flavanone isomerase (CHI) is an important enzyme of Phenylalaninemetabolic
pathways, in which the flavonoids are the end products. Flavonoids areinvolves in a variety of plant
biological processes, such has pigment formation,pheromone synthesis, response against pathogenic
microorganisms, legumenodulation and so on. The full-length peanut type CHI (CHI II) cDNA
andgenomic DNA were cloned in this study. The cDNA length of peanut CHI II was1184bp with an
ORF of675bp, encoding a peptide of224amino acids. The genomicsequence of peanut CHI
was2027bp in length containing four exons and threeintrons. Primers were designed accoding to the
coding sequence, and the promoterregion of CHIand CHI was cloned by chromosome walking.
Cis-elements ofthese promoters were analyzed using bioinformatic method. The results showed
thatCHI promoter was most probably root specific. Real-time quantitative PCR resultsconfirmed
the bioinformatic predicted result that CHI II was root specific. Theexpression of CHI II in different
peanut cultivars (with different color: green orpurpose in color) was measured. The results showed
that the expression of this genedid not correlated with the color of the peanut
plants.Totally,17homologous CHI genes from different plant species were downloadedfrom NCBI and
Phytozome Databases, and these sequences were used to analyse theevolution model of this gene
family use PAML software. The result showed that CHIgenes were under purifying selection during
evolution, and some amino acid sitesshowed neutral selection.In this study, different methods were
employed to anlysis the expressionregulation of BCCP and CHI gene families. The evolution of CHI

gene family wasdiscussed. This study provided valuable information for futher investigation aof
thesetwo gene families.