医学分子生物学杂志2012，Vol.9Issue(3)：204-207,4.DOI:10.3870/j.issn.1672-8009.2012.03.010
重组人胱抑素C的原核表达及鉴定
Prokaryotic Expression and Identification of Recombinant Human Cystatin C
张宏斌 1武婕 2赵华福 1王捷1
作者信息
- 1. 广州军区广州总医院医学实验科,广州市,510010
- 2. 广东省疾病预防控制中心病原微生物检验所,广州市,510600
- 折叠
摘要
Abstract
Objective To construct a prokaryotic expression vector of cystatin C (CysC) and purify recombinant Cys C protein produced by the expression system . Methods The gene of human CysC was obtained by synthesis. The cDNA fragment was cloned into prokaryotic expression vector pET-22b ( + ) to construct recombinant protein expression vector pET -Cys C. The positive recombinant clone was analyzed by restriction endonuclease digestion and DNA sequencing . The pET-Cys C plasmid was transfected into E. coli. BL 21 to induce CysC expression. The inclusion body protein was purified through Ni affinity chromatography and identified by using SDS -PAGE and Western blotting. Results The restriction enzyme digestion and DNA sequencing revealed the Cys C protein sequence coded by recombinant plasmid , completely corresponding to GenBank data. SDS-PAGE and Western blotting showed the size of expressed recombinant Cys C fusion protein at about 16kD. The purified recombinant Cys C through Ni affinity chromatography was greater than 90%. Conclusion In this study, we have established a high expression system of recombinant human Cys C protein.关键词
胱抑素C/原核表达/纯化Key words
cystatin C/prokaryotic expression/purification分类
医药卫生引用本文复制引用
张宏斌,武婕,赵华福,王捷..重组人胱抑素C的原核表达及鉴定[J].医学分子生物学杂志,2012,9(3):204-207,4.
