日本血吸虫(中国大陆株)谷胱甘肽转移酶编码区基因的体外扩增、克隆及在原核细胞的高效表达

为表达、获取日本血吸早谷胱甘肽转移酶（ＳｊＧＳＴ）基因工程重组蛋白，以日本血吸虫（中国大陆株）ｃＤＮＡ为模板，设计、合成特定寡核苷酸引物，ＲＴ－ＰＣＲ法扩增ＧＳＴ编码基因序列，将扩增产物连接ｐＧＥＭ－Ｔ克隆载体，再亚克隆到真、原核表达质粒ｐＢＫ－ＣＭＶ中，转染大肠杆菌ＸＬ１－ｂｌｕｅ，经ＩＰＴＧ诱导后用ＳＤＳ－ＰＡＧＥ分析表达效果。结果 ＲＴ－ＰＣＲ法特异性扩增出日本血吸虫ＧＳＴ编码区基因片段，其

Amplification,cloning and expression of the gene encoding 26KD Glutathione - S - Transferase from chinese Schistosoma Japonicum

To clone and expression of the gene encoding 26KD Glutathion-S-Transferase of Schistosoma japonicum(Chinese strain),Methods total RNA was isolated from adult worms using TRIzol reagent.The cDNA encoding 26KD Glutathion-S-Transferase was amplified by RT-PCR and cloned into pGEM-T vector,then,subcloned into the expressive plasmid vector pBK-CMV.The recombinants were identified by restriction enzyme analysis and PCR technique.The positive recombinant E.coli XL1-blue was induced by IPTG.SDS-polyacrylamide gel electrophoresis of total cellular protein from the bacteria was performed to detect the 26KD SjGST gene product.Results The objective gene was amplified and subcloned into expressive vector.The positive recombinant could be induced to express SjGST fusion protein with a molecular weight of 29KD.Induction 6 hours latter,the amount of exogenous proteins was about 30% of the total bacterial proteins.Conclusions The gene encoding 26KD SjGST was amplified and pBK-SjGST recombinants was successfully constructed.The recombinant 26KD SjGST protein was expressed in high efficiency and will be used to further study in immunodignosis and immunoprophyaxis of Schistomiasis japonica.