日本血吸虫中国大陆株基因重组抗原Sj22.6kDa基因克隆与表达的研究

目的 :分离 Sj22.6抗原基因 ,构建 Sj22.6表达克隆。方法 :抗体筛选 Sj成虫 cDNA库 ,PCR扩增目的基因片段 ,亚克隆入表达载体 pGEX- 1λt。对重组体进行诱导表达并对表达产物进行 SDS- PAGE和 Western blot分析。结果 :PCR扩增产物为约 930 bp的 DNA条带。亚克隆入pGEX- 1λt,获得两个表达克隆 pGSj6和 pGSj24 ,特异性表达产物为 22.6k Da抗原。重组体的表达方式为分离表达。结论 :从日本血吸虫成虫 cDNA库筛选到 Sj22 .6k Da克隆化基因 ,并构建了表达克隆 pGSj2 4和 pGSj6。

STUDIESON THE CLONING AND EXPRESSION OF THE RECOMBINANT Sj22 .6k Da ANTIGEN GENE OF SCHISTOSOMA JAPONICUM

AIM:To isolate Sj2 2 .6 gene and construct an expressive clone of r Sj2 2 .6 .METHODS: A S.japonicum adult c DNA library was screened by use of immune rabbit serum. From the possitive clones,one(λSj5 1 4,encoding Sj2 2 .6 ) was amplified by PCR to obtain its c DNA in- serts that was subcloned into the expressive plasmid vector p GEX- 1λtat Eco RI cloning site. The recombinants were induced by IPTG to express target protein which was analysed by SDS- PAGE and Western blotting. RESULTS:The specific product of PCR was about 930 bp which was subcloned in to pGEX-1λt. Two positive expressive clones were obtained, pGSj6 and pGSj24, which expressed specific protein with a molecular weigh t of 22.6 kDa. Th is recombinant protein ( rSj22.6) could be specifically recognized by immune rabbit serum. But the expressed product was not fusion protein, which meant that rSj22.6 was separated from the SjGST that is encoded by the plasmid pGEX-1λt. CONCLUSION: Sj22.6 antigen gene was screened from Sj adult cDNA library and subcloned in to pGEX-1λt, which resulted in two expressive clones, pGSj6 and pGSj24 that expressed rSj22.6.