马来丝虫肌球蛋白部分编码基因Bm-M55的克隆与真核表达  

Cloning and Eukaryotic Expression of the Gene Encoding Myosin from Brugia malayi

作　　者：谢东方[1] 方政[1] 黄为群[1] 沈勤[1] 童海燕[1] 徐邦生[1] 

机构地区：[1]南通大学医学院寄生虫学教研室,南通226001

出　　处：《中国寄生虫学与寄生虫病杂志》2008年第6期482-484,共3页Chinese Journal of Parasitology and Parasitic Diseases

基　　金：江苏省社会发展科技计划项目(No.BS2006522)~~

摘　　要：根据马来丝虫肌球蛋白部分编码基因(Bm-M55)序列设计引物,以其微丝蚴总RNA为模板,反转录PCR扩增目的基因。用TA克隆方法将目的基因克隆至载体pGEM-TEasy中,经PCR和双酶切鉴定并测序后,亚克隆至真核表达质粒pcDNA3.1(+),构建真核表达载体pcDNA3.1(+)/Bm-M55,转染COS-7细胞后进行RT-PCR验证。用十二烷基硫酸钠-聚丙烯酰胺电泳(SDS-PAGE)对获得重组蛋白Bm-M55进行分析和鉴定。RT-PCR鉴定结果显示,转染的COS-7细胞表达了Bm-M55基因,根据克隆的目的基因序列推导的氨基酸序列与GenBank(登录号为AAA27858)中的一致,重组蛋白Bm-M55相对分子质量(Mr)约为55000。Total RNA was extracted from periodic microfilariae of Brugia malayi and its myosin partial gene （Bm-M55） was amplified by RT-PCR. The PCR product was cloned and then subcloned into pcDNA3.1 （＋）vector. The recombinant eukaryotic plasmids were screened and identified by digestion with restriction enzyme and PCR amplification, and was transfected into COS-7 cells subsequently. The expressed protein was identified by SDS-PAGE. BIn-M55 mRNA was highly expressed in transfected COS-7 cells. The deduced amino acid sequence showed to be identical with that of Brn-M55, and the recombinant protein was about Mr 55 000.

关 键 词：马来丝虫 肌球蛋白 真核表达载体 COS-7细胞 

分 类 号：R383.161[医药卫生—医学寄生虫学]