GST-p16融合蛋白的表达、纯化及鉴定

Expression,purification and identification of recombinant GST-p16 fusion protein

目的 利用GST融合基因表达系统表达、纯化及鉴定GST p16融合蛋白。方法 以质粒pM p16为模板 ,扩增出p16基因片段 ,将其克隆至大肠杆菌表达载体pGEX 6P 1,将所构建的重组质粒pGEX p16转化大肠杆菌BL2 1并诱导表达 ,采用GST蛋白纯化系统进行纯化 ,所得产物进行SDS PAGE及Westernblot鉴定。结果 大肠杆菌细胞经诱导高效表达出约 4 2kD蛋白 ,其分子量与GST p16融合蛋白相符。Westernblot结果显示该蛋白能够被p16抗体特异性识别。结论 本实验在大肠杆菌表达系统中高效表达了有活性的GST

Objective To express,purify and identify recombinant GST p16 fusion protein using GST gene fusion system.Methods The cDNA of p16 was amplified from plasmid pM p16 and cloned into pGEX 6P 1 expression vector.The recombinant plasmid pGEX p16 was expressed in E.coli BL21 cells and the products were purified by GST purifying system.The specific expression was identified by Western blot.Results The resolved GST p16 fusion protein on 15% SDS PAGE showed a major band at position of 42kD and the fusion protein was recognized by anti p16 antibody on PVDF membrane.Conclusion The recombinant GST p16 fusion protein is expressed in E.coli cells efficiently,and the purified GST p16 fusion protein can be used in further study.