日本血吸虫32ku抗原基因的克隆及在耻垢分枝杆菌中的表达

Cloning and Expression of the Schistosoma Japonicum 32 ku Antigen Gene in Mycobacteria

日本血吸虫３２ ｋｕ抗原（简称ｓｊ３２ｋｕ）是血吸虫的一种保护性抗原，为研究其重组疫苗的保护作用，将该抗原基因的全编码序列按正确的阅读框顺序插入到穿梭质粒 ｐＢＣＧ２１００，受控于人结核杆菌热休克蛋白 ７０（Ｈｅａｔ ｓｈｏｃｋ ｐｒｏ－ｔｅｉｎ ７０，简称ｈｓｐ７０）启动子，构建成重组表达质粒 ｐＢＣＧ－ｓｊ３２，在大肠杆菌－分枝杆菌两个系统中能稳定复制。重组分枝杆菌于 ４２℃诱导 ２ ｈ或 ４５℃诱导 ３０ ｍｉｎ，经ＳＤＳ－ＰＡＧＥ，在 ３２ ｋｕ的位置能见到明显的蛋白条带，表明 ３２ ｋｕ抗原能在分枝杆菌中高效表达。

The Schistosoma japonicum 32 ku antigen (sj32ku) was a protective antigen in the adult. In order to construct the recombinant vaccine against Schistosoma japonicum, the complete coding sequence of sj32ku gene was inserted into the Escherichia coli-Mycobacteria shuttle plasmid (pBCG2100) which was controlled by M. tuberculosis heat shock protein70 (hsp70) protomer and could express foreign gene efficently. The recombinant plasmid (pBCG-sj32) was identified by PCR and restriction endonuclease cut. pBCG-sj32 could replicate in both E. coli and mycobacteria systems. The coding sequence of sj32 ku was cloned exactly into the downstream of the hsp70 promoter and could be expressed at 42℃, 45 ℃ in Mycobacteria. The expressed sj32ku protein could be observed in SDS-PAGE with the molecular weight of 32 ku.