重组大肠杆菌表达水稻白叶枯病菌FtsZ蛋白

摘要

旨在通过现代分子生物学技术制备水稻白叶枯病菌FtsZ蛋白。以水稻白叶枯病菌总DNA为模板，采用巢式PCR方法扩增获得水稻白叶枯病菌ftsZ基因，构建ftsZ基因的表达载体pET-22b-ftsZ，转化表达宿主E. coli BL21后，经PCR、NdeI/XhoI双酶切及测序鉴定、阳性克隆子经IPTG诱导表达，融合蛋白经镍柱纯化后，通过SDS-PAGE和Western blotting分析鉴定。结果显示，水稻白叶枯病菌ftsZ基因的重组表达载体构建成功，且阳性克隆子在IPTG的诱导下表达了FtsZ-6×His融合蛋白，并通过镍柱纯化获得了电泳纯的FtsZ-6×His融合蛋白。%It was to prepare FtsZ protein using techniques of modern molecular biology. TheftsZ gene was amplified fromXanthomonas oryzae by nested PCR, and recombinant plasmid pET-22b-ftsZ was constructed and transformed toE.coli BL21. The clony fragment was identificatified by PCR screening,NdeI/XhoI digestion and DNA sequencing, the positive clones were induced by IPTG for expression;the fusion protein was purified through Ni-NTA Resin, and identified by SDS-PAGE and Western blotting. Results showed that the recombinant plasmid pET-22b-ftsZ was constructed successfully, the FtsZ-6×His fusion protein was expressed in recombinedE. coli BL21 induced by IPTG, and purified through Ni-NTA Resin by electrophoretic purity.