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目的构建表达幽门螺杆菌(H.pylori)中性粒细胞激活蛋白A(NapA)的重组乳酸乳球菌菌株,为研究NapA作为疫苗抗原和免疫调节剂的应用效果奠定基础。方法用PCR方法从H.pylori基因组DNA中扩增napA基因,将napA经Nae I和SphⅠ双酶切与表达载体pNZ8110连接,用电穿孔法转入乳酸乳球菌菌株NZ3900中,并通过SDS-PAGE和Western blots分析鉴定napA表达产物。结果扩增的nap A基因长度为435 bp,构建的重组乳酸乳球菌经nisin诱导可表达分子量约为19 kD的重组蛋白,表达量约占菌体总蛋白量的12.4%,该重组蛋白可被小鼠抗H.pylori血清识别。结论成功构建了表达NapA的重组乳酸乳球菌菌株,该菌株在H.pylori口服疫苗和黏膜免疫调节研究中具有应用潜力。
Objective To construct a recombinant L. lactis strain expressing Helicobacter pylori neuratigenin A (NapA), which lays the foundation for the study of NapA as vaccine antigen and immunomodulator. Methods The napA gene was amplified from genomic DNA of H.pylori by PCR. The napA gene was ligated into expression vector pNZ8110 by Nae I and SphI digestion and transformed into Lactococcus lactis strain NZ3900 by electroporation. SDS - PAGE The napA expression product was identified by western blots analysis. Results The length of the amplified nap A gene was 435 bp. The constructed recombinant Lactococcus lactis was induced by nisin to express a recombinant protein with a molecular weight of about 19 kD, accounting for about 12.4% of the total bacterial biomass. The recombinant protein could be expressed Mouse anti-H. pylori serum recognition. Conclusion The constructed recombinant Lactococcus lactis expressing NapA has the potential application in oral vaccine and mucosal immunomodulation of H.pylori.