Studies On Molecular Basis Of The Regulation Of Trichome Formation In Tea Plant(Camellia Sinensis L.)

“Chahao” is the trichome on the abaxial surface of young leaf of tea plants(Camellia sinensis L.),which is closely related to the resistance of tea plant to environmental stress such as pests and diseases,and also affects the quality of tea.However,the regulation mechanism of the formation and development of trichome of tea leaf remains unclear.In this thesis,RNA-sequencing technology and quantitative PCR technology were used to analyze the dynamic changes of gene expression level in the leaves of two different cultivars of tea plants at different developmental stages,which contain rich or rare trichomes.Through enrichment analysis,transcription factor analysis and cluster analysis of differentially expressed genes(DEGs),candidate genes that might be involved in the development and regulation of trichome were screened.Moreover,the bioinformatic analysis and function prediction of Cs TT2 among the DEGs was carried out.Further,a plant overexpression vector of the CSTT2 candidate gene is constructed and the biological information and functional effects of the gene are constructed,and in order to further verify its function through heterologous expression.Finally,this thesis present work lays a foundation for revealing the molecular regulation mechanism of the formation of trichomes on tea leaves and the results were decribed as follwing:1.The results of RNA sequencing showed that 178.39 G of clean datae was obtained from six groups of samples,named as CYQ1,CYQ2,CYQ3,BDM1,BDM2,and BDM3,representing different amount of trichome on each leaf.This meaned that an average of more than 7.95 G clean data was obtained for each sample.Subsequently,a total of 48855 differentially expression genes were isolated,among which 16,271 genes were annotated as transcription factors,belonging to the 675 transcription factor familys.2.According to the related pathways of trichome development,351 DEGs were selected from the GO enrichment results,796 transcription factors were selected from the transcription factor family involved in the development of trichomes,and 2254 were selected through the VENN analysis of DEGs.Subsequently,the gene expression clustering and functional enrichment of these DEGs showed that the TEA004732 and TEA005642 genes may be involved in the regulation of the development of trichome of tea leaf.In addition,17 DEGs were obtained by screening genes related to trichome development among all DEGs.Among them,TEA031375 and TEA005642 genes expression level were consistent with the change trend of trichome of tea leaf,and they were considered to be involved in the development and regulation trichome in tea plant.3.The Cs TT2 was cloned by RT-PCR techniques and the plant overexpression vector p CXSN-Cs TT2 was constructed based on the vector p CXSN.The p CXSN-Cs TT2 was transferred into Agrobacterium GV3101,and then the Cs TT2 was transferred into Arabidopsis thaliana through the flower soaking using Agrobacterium GV3101 containing p CXSN-Cs TT2,which laid the foundation for the next step for functional identification.4.The bioinformatic analysis show that Cs TT2 encoded a protein with 294 amino acid residues which might be a hydrophilic protein,a non-secreted protein,and the protein was predicted to be located in the nucleus.The result of the tertiary structure indicated that Cs TT2 protein showed a 64.22% of identity with the structure of 6kks.1.A template,which is the typical structure of transcription factor WER family.According to the tertiary structure,Cs TT2 was predicted to be a kind of gene encoding the MYB transcription factor.In addition,the expression level of Cs TT2 was obsevered to be correlated with the amount of trichome on tea leaf.Therefore,Cs TT2 might be involved in the regulation of trichome development in tea plant.