鸡Ii基因的克隆、鉴定及其原核表达  

Cloning and Identifying of Chicken Invariant Chain Gene and Its Expression in Prokaryotic Cells

作　　者：刘玉华[1] 仲大莲[2] 许发芝[1] 余为一[1] 

机构地区：[1]安徽农业大学畜牧水产学院,合肥230036 [2]中国科学技术大学生命科学学院

出　　处：《安徽农业大学学报》2003年第4期390-394,共5页Journal of Anhui Agricultural University

基　　金：国家自然科学基金项目(30270974);安徽省"十五"科技攻关项目(01013003)资助。

摘　　要：从ConA诱导的5周龄SPF鸡脾细胞中提取总RNA,用自行设计的一对引物通过RT-PCR方法扩增出鸡恒定链(invariantchain,Ii)基因片段。经酶切鉴定后,再将该片段克隆到pcDNA3载体中,测定其DNA序列,结果表明该片段长度为669bp,其DNA序列与GenBank登录的基本一致。再将该片段插入原核表达载体PGEX-4T-1,并导入菌株BL21,经诱导培养、蛋白提取和SDS-PAGE,获得分子量约为50kD的特定蛋白条带,表明所克隆的鸡Ii基因在原核细胞中得到表达。A total RNA was obtained from 5 weeks old SPF chicken spleen cells stimulated by ConA, then cDNA of invariant chain (Ii) gene was cloned from the RNA by reverse transcription (RT)-PCR with a pair of designed primers.The length of this cloned fragment was 669 bp and was identified by a restriction endonuclease cleavage.For sequencing,it was further inserted into plasmid pcDNA3,and showed identical with the chicken Ii gene registered in the GenBank. This fragment was inserted into PGEX-4T-1, a prokaryotic expressive plasmid, and the latter was transferred into the strain BL21. After the bacteria were induced and cultured, expressed protein was separated. A band with 50 000 of molecular weight was observed on SDS-PAGE. All these results suggest that the cloned gene of chicken Ii could be expressed in prokaryotic cells.

关 键 词：鸡 II基因 克隆 原核表达 

分 类 号：S852.4[农业科学—基础兽医学]