原核表达重组质粒pUC57-CTP-PTEN的构建及鉴定

Construction and identification of the prokaryotic expression recombinant plasmid pUC57-CTP-PTEN

目的:合成基因CTP及PTEN基因,构建其原核表达重组质粒pUC57-CTP-PTEN,为后续研究其功能做准备。方法:合成基因CTP-PTEN,将该基因定向克隆到原核表达载体pUC57中,构建pUC57-CTP-PTEN重组质粒,转化大肠杆菌DH5α菌株,抽提质粒进行双酶切、测序鉴定后,用Western blotting检验表达。结果:测序鉴定CTP-PTEN基因合成成功。经双酶切、测序鉴定证实重组质粒pUC57-CTP-PTEN成功转入DH5α,Western blotting检验pUC57-CTP-PTEN成功表达。结论:成功构建了重组质粒pUC57-CTP-PTEN,并在大肠杆菌DH5α中成功表达。

Objective： To construct prokaryotic expression recombinant plasmid p UC57- CTP- PTEN of CTP gene and PTEN gene,and prepare for the follow- up study of its function. Methods： Gene CTP- PTEN was synthesized and cloned into the prokaryotic expression vector p UC57 to construct the recombinant plasmid p UC57- CTP-PTEN. Then it was transformed into E. coli DH5 α. The recombinant plasmid was identified by restriction enzyme digestion and DNA sequencing. The expression product was identified by Western blotting assay. Results： Gene CTP-PTEN was successfully synthesized. The recombinant plasmid p UC57- CTP- PTEN was identified that it was successfully transformed into E. coli DH5 α and expressed by restriction enzyme digestion,DNA sequencing and Western blotting. Conclusion： The recombinant plasmid p UC57- CTP- PTEN was successfully constructed and expressed in E. coli DH5 α.