采用SMART RACE技术成功克隆了安吉白茶茶氨酸合成酶基因（TS）的全长cDNA序列,并将其登录Genbank,登录号为JN226569。TS基因的cDNA全长为1503bp,开放阅读框（ORF）为1071bp,编码356个氨基酸。经生物信息学分析,TS蛋白的氨基酸残基数为356,分子量为39250.3Da,理论等电点（PI）为5.79,其氨基酸序列中可能不存在卷曲螺旋结构。TS蛋白不存在信号肽序列,不发生跨膜运动,属于亲水性的非分泌蛋白,其磷酸化位点有26个。通过亚细胞定位预测,安吉白茶TS蛋白是一种细胞质蛋白。克隆茶氨酸合成酶基因的全长cDNA序列,对于利用生物技术手段改良茶树品种,以调控茶树中茶氨酸的含量具有重要意义。
The full-length cDNA of theanine synthetase (TS, GenBank accession No.JN226569) gene of Anji white tea was cloned. The cDNA sequence has the full-length of 1503bp, and with an Open Reading Frame of 1071bp. It can encode 356 amino acids. Bioinformatics analysis showed that theanine synthetase has 356 amino acid residues, has a theoretical molecular weight of 39250.3Da and theoretical PI value of 5.79. Bioinformatics prediction showed that this protein is hydrophilic and located not within the transmembrane. There are 26 phosphorylation sites within the polypeptied chain. The signal peptide analysis showed that there is no signal peptide and no winded helix domain present within the sequence, the protein is non-secreted and it functions in the cytoplasm. This reported result about cloning full-length cDNA of theanine synthetase gene would be beneficial for using biotechnology to improve tea plant cultivar resources and regulate L-theanine level.
[1] 张莹, 施兆鹏, 施玲. 茶氨酸的研究进展[J]. 天然产物研究与开发, 2003, 15(4): 369-372.
[2] Arai M, Matsuura T, Kakuda T.Variation of the yields and the chemical composition in leaves on nutriculture of tea plant[J]. Nippon Dojo Hiryogaku Zasshi, 1989, 60: 157-159.
[3] 汤皴. 茶氨酸的合成药理及其在食品中的应用[J]. 茶叶通报, 2002, 24(4): 19-21.
[4] 陈宗懋. 茶氨酸具有降压功能[J]. 中国茶叶, 1997, 19(2): 27.
[5] 丁王, 丁雪. 茶氨酸的抗疲劳作用研究[J]. 中国公共卫生, 2002, 18(3): 315-317.
[6] Sasaoka K, Kito M, Y O, et al. Some properties of the theanine synthesizing enzyme in tea seedlings[J]. Bio Chem, 1965, 29(11): 984-988.
[7] 李娟, 刘硕谦, 刘仲华, 等. 一种改良的茶树高质量RNA快速提取方法[J]. 福建茶叶, 2007(4): 34-35.
[8] 董志敏, 李英慧, 张宝石, 等. 一种获得大片段克隆的SMART全长cDNA文库构建方法[J]. 大豆科学, 2007, 12(3): 223-226.
[9] 王朝霞, 李叶云, 江昌俊, 等. 茶树巯基蛋白酶抑制剂基因的cDNA克隆与序列分析[J]. 茶叶科学, 2005, 25(3): l77-182.
[10] Mizutani M, Nakanishi H, Ema J, et al. Cloning of β-primeverosidase from tea Leaves, a key enzyme in tea aroma formation[J]. Plant Physiology, 2002, 130: 2164-2176.
[11] 邓婷婷. 安吉白茶白期叶与绿期叶差异表达基因的全长cDNA克隆与原核表达[D]. 长沙: 湖南农业大学, 2009.
[12] Ye Z, Connor JR. cDNA cloning by amplification of circularized first strand cDNAs reveals non-IRE-regulated iron-responsive mRNAs[J]. Biochem Biophys Res Commun, 2000, 275(1): 223-227.
[13] 董志敏, 李英慧, 张宝石, 等. 一种获得大片段克隆的SMART全长cDNA文库构建方法[J]. 大豆科学, 2007, 12(3): 223-226.
浙公网安备 33019902000101号 
