摘要
目的:构建结核分枝杆菌Rv1884c基因的原核表达质粒,获得结核分枝杆菌Rv1884c基因的表达蛋白。方法:制备结核分枝杆菌基因组DNA,采用聚合酶链反应技术扩增目的基因片段;通过pGEX-4T-1构建质粒载体pGEX-4T-1-Rv1884c,经序列测定证实正确后转化大肠杆菌DH5α,再经IPTG诱导表达GST-1884融合蛋白;用聚丙烯酰胺凝胶电泳分析重组蛋白的相对分子质量及表达形式。结果:扩增出了结核分枝杆菌Rv1884c基因,构建了具有正确基因序列的质粒载体pGEX-4T-1-Rv1884c,转化大肠杆菌DH5α后经诱导产生了高水平的表达产物。结论:构建了pGEX-4T-1-Rv1884c质粒载体,并诱导表达了GST-1884融合蛋白,为进一步研究Rv1884c蛋白的活性及其功能,探讨结核分枝杆菌快速促生长作用奠定了基础。
Objective： To construct fusion gene and prokaryotie expression plasmid of the Rv1884c gene of Mycobacteriurn tuberculosis, and to express the fusion proteins efficiently in Escherichia coll. Methods： The Rv1884c gene was amplified by PCR with specific primers from genomic DNA of M.tuberculosis H37Rv strain, and was cloned into pGEX-4T-1 expression vector. After sequenced, DHSa strain of E.coli was transformed with the recombinant vector and induced to express recombinant proteins. The relative moleculer size of the proteins were analyzed by SDS-PAGE. Results： The length of PCR products was 531 bp and identical with what the GenBank reported. The recombinant expressive vector pGEX4T-1-Rv1884c was constructed. The E.coli DHSa strains with recombinant vector showed high level of Rv1884c gene expressions after IPTG induction. Conclusion： The fusion Rv1884c gene and prokaryotic expression plasmid were constructed. The expression of recombinant Rv1884c protein with natural activity lays a basis for further study of fast cultivation in M. tub erc ulos is.
出处
《生物技术通讯》
CAS
2007年第2期202-204,共3页
Letters in Biotechnology
