Prokaryoti Expression,Purification and Activity Analysis of Human Adiponectin Gene

Objective: To express recombinant human adiponectin with biological activity in E coli. Methods: The cDNA coding fragment of human adiponectin gene was amplified by PCR. Gateway prokaryoti expression system was used. PCR products were cloned into pET-DEST42 plasmid to construct fusion expression vector pET-DEST42/attB-adiponectin after BP reaction and LR reaction. pET-DEST42/attB-adiponectin was transformed into E coli BL21 and the bacteria were induced by IPTG expressed adiponectin,fusion protein was purified by Ni·NTA affinity chromatography. The effect of purified recombinatant adiponectin on glucose-6-phosphatase was detected by RT-PCR. Results: The DNA sequencing showed that expression vector pET-DEST42/attB-adiponectin was constructed successfully. Purified adiponectin showed antigenicity and reduced glucose-6-phosphatase transcriptional level. Conclusion: We have obtained the recombinant human adiponectin with biological activity,which is significant for further study.