摘要
试验以构建的海南黄牛外周血白细胞cDNA文库为材料,采用菌落PCR的方法筛选出海南黄牛细胞分裂周期蛋白42(cell division cycle 42,Cdc42)全长cDNA,构建pET28a-Cdc42重组质粒,经IPTG诱导表达后,进行可溶性分析和蛋白纯化,并应用Western blotting对其进行鉴定。结果显示,海南黄牛Cdc42 cDNA的编码框由576个碱基组成,编码191个氨基酸,融合蛋白分子质量约为30 ku,该蛋白主要以包涵体形式存在;纯化后的蛋白经Western blotting检测,发现对应大小的特异性条带,表明本试验成功克隆Cdc42并表达。
Hainan Yellow cattle peripheral blood leukocyte cDNA library was used to screen Cdc42 cDNA and pET28a-Cdc42 recombinant plasmid was constructed.Protein expression was induced by IPTG,and analyzed by SDS-PAGE and Western blotting.The results showed that the Cdc42 was composed of 576 bp,encoding 191 amino acids;the protein about 30 ku as expected was expressed after induction with IPTG;after purification,the corresponding protein strip was detected by Western blotting.The data indicated that prokaryotic expression vector of Cdc42 was constructed and expressed successfully.
出处
《中国畜牧兽医》
CAS
北大核心
2011年第6期68-71,共4页
China Animal Husbandry & Veterinary Medicine
基金
国家高技术研究发展计划(863)项目(2011AA100302)
海南省重点科技项目(06102)
关键词
海南黄牛
CDC42
原核表达
纯化
Hainan Yellow cattle
Cdc42
prokaryotic expression
identification
