中国黄牛PrP^c成熟蛋白基因的克隆和序列分析

Cloning and Sequencing of PrP^c Mature Protein Gene of Chinese Yellow Cattle

根据已发表的牛PrPc蛋白基因序列 ,设计合成一对引物 ,并在2个引物的两端分别加入BamHI和NdeI酶切位点。以从中国黄牛肝脏中提取的基因组DNA为模板 ,经PCR扩增出一约640bp的目的基因片段 ,将此基因克隆于 pET11a载体 ,构建了PrPc蛋白基因重组质粒b-pET11a -PrPc ,转化DH -5α并进行序列测定。同源性分析表明 :中国黄牛PrPc成熟蛋白基因与目前国外发表牛的成熟蛋白PrPc基因相比 ,有较大差异 ,发现了一个新的NdeI酶切位点 ;推导的氨基酸序列有4个氨基酸差异 。

A pair of primers were designed and synthesized based on the publised bovine PrPc protein gene sequense and Bam HI and Nde were added to the two ends of the primers respectively. A target gene fragment about 640 bp was amplified by PCR using the genosome DNA extracted from Chinese yellow cattle liver as template and cloned into plasmid pETlla , thus constructing PrPc protein gene recombinant plasmid b-pETlla-PrPc, transformed into DH-5a and sequenced. Homegenetic analysis showed that there are considerable differences between PrPc mature protein gene of Chinese yellow cattle and the published bovine PrPc mature protein gene abroad: a new Nde I restriction site was found; there were four amino acids variation in the derived amino acid sequences and lacked an eight amino acid repeated zone.