Construction of prokaryotic expression vector of rhLEDGFp52 gene、 inducing expression and purification of its protein
- VernacularTitle:rhLEDGF p52基因原核表达载体的构建、诱导表达及蛋白纯化
- Author:
Hai-Sheng, ZHAO
;
Yi, WANG
- Publication Type:Journal Article
- Keywords:
rhLEDGFp52;
gene expression;
E.coli;
protein purification
- From:
International Eye Science
2006;6(4):751-754
- CountryChina
- Language:Chinese
-
Abstract:
AIM: To construct the prokaryotic expression vector of rhLEDGFp52 gene,to obtain the rhLEDGF p52 protein.METHODS: rhLEDGFp52 gene was constructed into a prokaryotic expression vector pET30a (+) by recombinant DNA techniques and was identified by enzymatic digestion and sequence analysis. rhLEDGFp52 protein was induced expression by IPTG in E.coli BL21 (DE3), it was tested by Western blot and was purified by Ni-NTA His. Bind. Resin.RESULTS: We Successfully constructed the prokaryotic expression vector of rhLEDGFp52 gene and obtained its expression in E.coli BL21 (DE3),it was expressed in a soluble form and detected up to 34.63% of the total bacterial protein expressed in E.coli BL21 (DE3).Western blot analysis demonstrated that rhLEDGFp52 protein could spicifically integrate with LEDGF-ab. After purified by Ni-NTA His. Bind. Resin, the ultimate concentration of purified rhLEDGFp52 protein was 520μ g/ml and its purity was 87.93%.CONCLUSIONS: rhLEDGF p52 protein was obtained that provides an experimental basis for the further study of the biological function of rhLEDGFp52 protein.
