[Cloning of the glyceraldehydes 3-phosphate dehydrogenase gene of porphyromonas gingivalis and its expression in E. coli].

作者：Ang Li, Hongyan Xu, Jian-feng Shi, Chunhui Zhu, Guo-zhou Rao, Jianzhong Gou · 发表于：PubMed · 年份：2011 · 被引用次数：2 · 研究领域：Molecular Biology Techniques and Applications、Oral microbiology and periodontitis research、Enzyme Production and Characterization

OBJECTIVE: To clone the glyceraldehydes 3-phosphate dehydrogenase (GAPDH) gene of Porphyromonas gingivalis (P. gingivalis) and to induce its fusion expression in E. coli. METHODS: GAPDH was obtained by PCR and was inserted into cloning vector pMD-18-T to construct clone recon. Double enzymes digest the recon pMD18-T-GAPDH and the prokaryotic expression vector pET-32a and then connect to get the expressing recon pET-32a-GAPDH. The recombinant expression plasmid which had been confirmed by enzymes digestion was transformed to E. coli competent cells BL21 and induced the expression of GAPDH with isopropyl beta-D-1-thiogalactopyranoside (IPTG) of different density. RESULTS: DNA sequencing showed that the fragment was 99.802% the same to the sequence published in NCBI. Under the best density, IPTG could be highly expressed. CONCLUSION: The GAPDH had been successfully cloned and expressed in E. coli which gets ready for the following experiment to study the immunity of GAPDH and the homologues antibody preparation.