柳蚕S3a基因的鉴定及原核表达分析

Identification and Prokaryotic Expression Analysis of S3a from Actias selene

采用RT-PCR法克隆了柳蚕S3a基因序列,并通过构建重组质粒,对其进行了原核表达及蛋白纯化。柳蚕S3a基因ORF长度为795 bp,编码264个氨基酸,预测蛋白的等电点和分子量大小分别为9.74和30 kD。序列比对及进化关系表明,柳蚕S3a蛋白与其他物种S3a蛋白相似度介于71%和97%之间,其中与鳞翅目昆虫的同源性最高。SDS-PAGE和Western blotting结果显示柳蚕S3a在大肠杆菌中获得了高效表达,分离获得的重组蛋白的纯度较高。

RT-PCR was used to identify S3a gene from Actias selene and prokaryotic expression and purification of S3a were also performed by construction of recombinant plasmids. An ORF consisted of 795 bp encoding a protein with 264 amino acids was identified in A. selene S3a and predicted pl and MW were 9.74 and 30 kD for this protein, respectively. The sequence comparison of S3a proteins between A. selene and other animals was carried out, and the similarity was ranged from 71% to 97 % with higher homologue to Lepidoptera insects. The results of SDS-PAGE and Western blotting showed that recombinant proteins were efficiently expressed in E. coli and pure recombinant proteins were successfully acquired.