幽门螺杆菌热休克蛋白A基因的克隆及论著高效分泌表达

Cloning and high secretion expression of heat-shock protein A gene from Helicobacter pylori

目的 :从幽门螺杆菌 (Hp)分离热休克蛋白A基因 ,并实现在大肠杆菌中的融合分泌表达。方法 :提取Hp染色体DNA ,用PCR方法扩增hspA基因。经克隆、测序后 ,构建融合分泌表达载体 pMAL HspA ,转化大肠杆菌 ,IPTG诱导表达。 结果 :分离得到了高度保守的 354bp的hspA基因片段 ,并在大肠杆菌中实现了该基因的融合分泌表达。在 37℃诱导表达 3h后 ,其融合分泌表达蛋白可占细菌周质总蛋白的 66.6%。该表达带可被抗Hp的特异抗体所识别。 结论 :幽门螺杆菌hspA基因的克隆与表达为Hp疫苗的研制打下了基础。

Objective: To isolate and express the heat-shock protein A(hspA) gene from Helicobacter pylori(Hp). Methods: The hspA gene was amplified from Hp chromosomal DNA by PCR. After sequencing, the hspA gene was recombined in vitro with fusion secretion expression vector pMAL-p2 and transformed into E.coli cells. Results:The highly conservative 354 bp hspA gene fragments were isolated. Recombinant E.coli strains expressing HspA were successfully obtained. Fusion expression protein amounted to 66.6% of the total bacterial periplasm protein after induced with IPTG for 3 h at 37℃. The expression product could react specifically on anti-Hp rabbit IgG. Conclusions: Cloning and high-expression of hspA gene from Hp provided a basis to construct the Hp vaccine. [