Cloning, expression and purification of human MRPS17 cDNA in Escherichia coli / 解放军医学杂志

ABSTRACT

Objective To clone, express and purificate human MRPS17 cDNA in Escherichia coli (E. coli). Methods MRPS17 cDNA was obtained from total RNA isolated from primary cultured human hair papillary cells by RT-PCR and sequencing method, and then it was inserted into prokaryotic expression vector pET28a for the IPTG-induced expression in E. coli BL21 (DE3). The expression product fused with 6?His at C-terminal was analyzed by Western blotting, and purified by using Ni 2+-NTA ion exchange resin. The purity of MRPS17 protein was analyzed by SDS-PAGE. Results Human MRPS17 cDNA was obtained, and the expression plasmid pET28a-MRPS17 was constructed successfully. Western blot analysis showed that human MRPS17 with 13kD molecular weight was expressed in E. coli at 20℃. The purity of the recombinant MRPS17 was more than 90% after purification using Ni 2+-2NTA ion exchange resin. Conclusion The sequence of MRPS17 cDNA was consistent with the known sequence from the Genbank. MRPS17 is successfully induced and expressed in E. coli.