诸葛菜(Orychophragmus violaceus)Toc33基因编码区的克隆及序列分析  

Molecular Cloning and Sequence Analysis of Orychophragmus violaceus Toc 33

作　　者：廖问陶[1] 赵志伟[1] 王茂林[1] 赵云[1] 

机构地区：[1]四川大学生命科学学院,成都610064

出　　处：《四川大学学报（自然科学版）》2003年第1期163-167,共5页Journal of Sichuan University(Natural Science Edition)

基　　金：国家自然科学基金(30170500)

摘　　要：以诸葛菜(Orychophragmusviolaceus)新鲜嫩叶为材料提取总DNA,并以其为模板,根据已报道的拟南芥(Arabidopsisthaliana)Toc33基因的编码序列,设计一对PCR引物,扩增诸葛菜叶绿体外膜蛋白转运器构件蛋白基因Toc33,得到两条扩增带,将这两条带分别割胶回收,与T载体连接转化E.coli,筛选重组子并测序,结果表明克隆到的这两个片段分别长1475bp,1573bp,通过同源性比较发现,它们之间的同源性达到88%.这两个片段与拟南芥Toc33基因有较高的同源性,分别命名为OvToc33 1,OvToc33 2.参考拟南芥Toc33外显子和氨基酸序列,分别确定了所克隆的两个诸葛菜Toc33基因片段的7个外显子和6个内含子,得到了诸葛菜Toc33基因的全编码区序列,并推导了其氨基酸序列.这两个DNA序列与拟南芥Toc33基因编码区的同源性分别高达91.8%和92.4%,说明这一蛋白是高度保守的.By using PCR with primers designed according to the Toc33 sequence of Arabidopsis thaliana, two homologous DNA sequences were directly amplified from the genomic DNA of Orychophragmus violaceus. The two PCR fragments were cloned and confirmed by PCR and restriction enzyme digestion. Sequence analysis showed that the two sequences were1475bp, 1573bp and shared 88% identity with each other, their coding region and deduced amino acid sequences also showed very high identity with each other and with Arabidopsis thaliana Toc33 sequence, the result showed that the sequences are highly conserved.

关 键 词：编码区 诸葛菜 Toc33基因 序列分析 叶绿体 蛋白转运 基因克隆 

分 类 号：Q785[生物学—分子生物学]