Construction of a prokaryotic expression vector of PHD2 and its expression in E. coli

Objective To achieve soluble expression of Nus-PHD2 fusion protein in E. coli. Methods pET-43. 1b( +) vector was digested by SacI and the linearized vector was purified. PHD2 gene was amplified from pCMV6-Entry-EGLN1 plasmid. Specific In-Fusion PCR primers were designed to make the PCR products contain ends that were homologous to the linearized vector. PHD2 gene was cloned into the linearized pET-43. 1b( +) vector using In-Fusion technology. The recombinant expression plasmid was then transformed into BL21( DE3) and induced to express the Nus-PHD2 fusion protein. The recombinant protein was analyzed and identified by SDS-PAGE and Western blot. And the target protein was purified by Ni-NTA His·Bind. Results The prokaryotic expression plasmid pET-43. 1b( +)-PHD2 was successfully constructed. The recombinant protein was solubly expressed and could specifically react with anti-PHD2 antibody. Conclusion Soluble expression of Nus-PHD2 fusion protein was achieved in E. coli,which lay a foundation for further study on the biological functions of PHD2.