Escherichia coli expression of lactate dehydrogenase C gene of black-lipped pika in Western Sichuan Plateau.

[Objective] To express lactate dehydrogenase C (LDH-C) gene in prokaryotic system and then purify the recombinant protein. [Method] The LDH-C gene was amplified from black-lipped pika (Ochotona curzoniae) by RT-PCR and ligated to the expression vector pET-32a. The E. coli BL21 (DE3) carrying the LDH-C gene was obtained and induced by IPTG (isopropy-beta-D-thiogalactoside). The expression products were analyzed by SDS-PAGE and purified by affinity chromatography. [Result] An approximately 1.0 kbp band appeared in the RT-PCR products as theoretically expected. As evidenced by PCR and double enzyme digestion, an approximately 1.0 kbp fragment appeared, which indicated successful construction of expression vector. As analyzed by SDS-PAGE, a fusion protein with molecular weight slightly larger than 45 kDa was expressed in form of inclusion body. And very pure fusion protein was obtained by nickel ions-charged affinity chromatography. [Conclusion] The LDH-C gene of black-lipped pika has been cloned and expressed in E. coli.