摘要
目的:构建食管癌相关基因2(ECRG2)原核表达质粒,在大肠杆菌(E.coli)中诱导表达重组蛋白。方法:用聚合酶链反应法扩增ECRG2基因开放阅读框(ORF),构建表达载体pGEX-4T-1-ECRG2,测序鉴定后转化E.coli(BL21)进行异丙基-β-D-硫代半乳糖苷诱导表达,目的蛋白进行SDS-聚丙烯酰胺凝胶电泳(PAGE)、Western免疫印迹鉴定。结果:SDS-PAGE测定GST/ECRG2蛋白分子质量约34 ku,Western blot验证出目的蛋白特异表达。结论:ECRG2 ORF插入pGEX-4T-1质粒并在E.coli中成功表达。
Objective： To construct a prokaryotic expression vector containing esophageal cancer related gene 2 （ECRG2）and observe its expression in Escherichia coli（E.coli）. Methods： Open read frame of ECRG2 was amplified by polymerase chain reaction and linked into vector pGEX-4T-1 by T4 DNA ligase. Then the recombined expression plasmid was transduced into BL21 by TSS, and induced by 1 mmol/L IPTG at 37 ℃. Results： The fusion protein GST/ ECRG2 about 34 ku was detected by SDS-PAGE and Western blot after lysis of bacteria. Conclusion： A prokaryotic expression plasmid containing ECRG2 gene is successfully constructed, which can highly express objective protein in E. coli .
出处
《汕头大学医学院学报》
2006年第3期143-145,共3页
Journal of Shantou University Medical College
基金
国家基础研究重点项目(973)(2002CB513101
2004CB518701)
博士后基金资助
关键词
食管癌相关基因2
基因克隆
原核表达
esophageal cancer related gene 2
gene clone
gene expression
