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目的:构建日本血吸虫单克隆抗独特型抗体NP30重链可变区6×CDR3基因/IL-2融合基因,以观察其对动物的抗血吸虫感染保护作用。方法:通过PCR方法体外扩增NP30 6×V_HCDR3和IL-2基因,经测序后先后重组入原核表达质粒pET-28a(+)相应的位点上,将构建正确的6×V_HCDR3/IL-2-pET-28a(+)表达质粒转化E.coli BL21,IPTG诱导表达。结果:构建的6×V_HCDR3/IL-2融合基因中,6×V_HCDR3和IL-2序列均正确,融合基因经IPTG诱导表达重组蛋白,十二烷基磺酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)分析分子质量约为30ku,表达量约占菌体总蛋白的20％。结论:成功构建并原核表达了NP30 6×V_HCDR3/IL-2融合基因。
Objective: To construct the 6 × CDR3 gene / IL-2 fusion gene of Schistosoma japonicum monoclonal anti-idiotypic antibody NP30 heavy chain variable region to observe the protective effect against schistosoma infection in animals. Methods: The NP30 6 × V_HCDR3 and IL-2 genes were amplified by PCR in vitro and sequenced. The recombinant plasmids were recombined into the corresponding sites of the prokaryotic expression plasmid pET-28a (+) to construct the correct 6 × V_HCDR3 / IL-2 -pET-28a (+) expression plasmid was transformed into E.coli BL21 and induced by IPTG. Results: The sequences of 6 × V_HCDR3 and IL-2 were all correct in the constructed 6 × V_HCDR3 / IL-2 fusion gene. The fusion gene was induced by IPTG to express recombinant protein, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis SDS-PAGE) analysis of molecular weight of about 30ku, the expression of about 20% of the total bacterial protein. Conclusion: NP30 6 × V_HCDR3 / IL-2 fusion gene was successfully constructed and prokaryotic expressed.