甘蓝型油菜Toc33基因编码区的分离及蛋白序列的比较

Molecular cloning and sequence analysis of Toc33 gene in Brassica napus

以甘蓝型油菜新鲜嫩叶为实验材料提取其总DNA,以其为模板,根据拟南芥Toc33基因编码区序列设计引物,PCR扩增甘蓝型油菜叶绿体外膜蛋白转运机器的构件蛋白基因Toc33,得到两条扩增带,测序结果显示克隆到的两个片段分别长1370bp、1490bp,将这两个片段分别命名为BnToc33-1,BnToc33-2,序列比较发现它们之间的同源性为78%,其中外显子的同源性为96%,而内含子的同源性仅为60%.为研究Toc33与同一基因家族的Toc34基因功能间的关系,对拟南芥、油菜、诸葛菜等植物的Toc33、Toc34蛋白序列进行比较分析并构建了分子系统进化树.

By using primers according to the Toc33 sequence of Arabidopsis thaliana, two homologous DNA sequences were amplified from the genomic DNA of Brassica napus. Sequence analysis showed that the two sequences were 1 370 bp and 1 490 bp respectively; they shared 78% identity with each other, their coding region and deduced amino acid sequences also showed very high identity with Arabidopsis thaliana Toc33 sequence. Based the alignment of the amino acids of Toc33 and Toc34 genes, the evolutionary tree of Brassica napus, Zea mays, Pisum sativum, Arabidopsis thaliana and Orychophragmus violaceus was constructed.