幽门螺杆菌NCTC11639基因hpaA的克隆表达及鉴定

Cloning and Characterization of Helicobacter Pylori Gene HpaA from Strain NCTC11639

目的获取幽门螺杆菌(Hp)hpaA基因的序列,预测其编码蛋白hpaA作为候选疫苗的可行性,在大肠杆菌表达系统中表达hpaA。方法提取Hp标准株NCTC11639的基因组DNA,使用PCR扩增hpaA基因,克隆入pGEM-T载体中测序,并进行同源性分析。将hpaA克隆入表达载体pGEX-4T-1中,诱导表达,SDS-PAGE电泳鉴定。结果同源性分析表明hpaA具有很高的保守性,成功构建出可以高效、分泌型表达hpaA的原核表达系统。结论hpaA具有高保守性,在Hp各菌株中普遍存在,是有良好应用前景的Hp候选疫苗。

Objective The intention of this study can be subscribed in three aspects： obtaining the sequence of Helicobacter pylori gene hpaA ,predicting whether protein hpaA has potential to become a vaccine candidate of Hp and expressing hpaA in E.coli heterogeneous expression system. Methods The hpaA gene was amplified from the genome of Hp strain NCTC11639 and then cloned into vector pGEM-T. The sequence has been obtained by sequencing the recombinant vector. The pGEX-4T-1 was used to construct expression system in E. Coli strain BL2.1 and the expression result verified by SDS-PAGE. Results The homogeneous analysis showed that hpaA is a highly conservative gene. An efficient and secretive expression system in BL21 was constructed. Conclusion The research indicates that hpaA is a universaal and highly conservative gene and hpaA is a promising vaccine candidate of Helicobacter pylori.