Differential Gene Expression In Tea (Camellia Sinensis L.) Calluses With Different Catechins Contents By CDNA-AFLP

Catechins are the most important featured metabolic components of tea leaves and also main chemical compositions contributing to the tea quality. Studies on gene expressions in the catechins biosynthesis are of great practical significance for improving breeds of tea plants and tea quality.By applying cDNA-AFLP, this study created the cDNA-AFLP profile of calluses with different catechins concentrations ("Yunjing 63X"and"Yunjing 63Y"), analyzed differential gene expressions of experiment materials; by using qRT-PCR, expression differences of relevant genes in catechins biosynthesis were elaborated; RACE was adopted to clone a MYB gene of tea plant which might influence the growth of cells. Following conclusions were obtained.From the perspective of appearances and growth conditions,"Yunjing 63Y"grew faster than"Yunjing 63X", and the former appeared yellowish with relatively harder texture while the latter appeared whitish with relatively looser texture. HPLC results showed compositions of catechins in"Yunjing 63X"and"Yunjing 63Y"differed slightly, and both of them contained the basic five catechins, that is, C, EC, EGC, EGCG and ECG, but the contents of all these single catechins and the total catechins in Yunjing 63Y were higher than those in"Yunjing 63X".cDNA-AFLP was used to identify differentially expressed genes in both of the calluses, 60 different expressed sequence tags (EST) with lengths between 110-1000bp were screened, the expression of 41 of them was higher in"Yunjing 63Y"while 19 in"Yunjing 63X". Blast comparisons indicated that all ESTs could be classified into 7 putative cellular functions: phyenylpropanoid metabolism (2), UDPG-dependant transferase (5), transcription factor (8), transporter (7), signal transduction (10), other metabolisms (17) and unclear functions (11).With calluses and fresh tea leaves as experiment materials, qRT-PCR was applied to analyze the different expressions of dominant predicted genes in cDNA-AFLP profile (phenylalanine ammonialyase, UDPG: anthocyanin 3'-O-glucosyltransferase, CsMYB1, CsMYB2,WD40, 2,4-D inducible glutathione-S-transferase) and relevant genes of catechins biosynthesis (PAL, F3H, DFR, LAR, ANS and ANR). The results showed that the expression of all these genes was higher in"Yunjing 63Y", and with the changing maturity of fresh leaves, the expression laws of these genes also showed differences.RACE was applied to amplify the full-length cDNA of CsMYB1 which comprised 1270bp cDNA with an ORF of 933bp, encoding 310 amino acids, the predicted molecular weight 105.27 KDa, isoelectric point 4.85. Blast comparisons also showed that the full-length cDNA of CsMYB1 shared high homology with MYB protein of many plants, thus it was predicted that CsMYB1 is a MYB proteoid gene relevant to transcription factor in tea plant, which might influence stress signal.