结核分支杆菌Ag85B基因真核表达株的构建(英文)

Construction of eukaryotic expression strain encoding Ag85B gene of mycobacterium tuberculosis

目的 构建结核分支杆菌分泌蛋白Ag85B编码基因的DNA疫苗。方法 以结核分支杆菌H37Rv基因组为模板 ,PCR扩增Ag85B编码基因 ,用限制性内切酶消化后 ,插入真核表达载体pVax1中 ,转化大肠杆菌DH5α ,进行重组子筛选、鉴定。结果 以结核分支杆菌PCR扩增为阳性 ,经限制性内切酶消化后可见目的条带 ,所测定序列与报道的Ag85B序列一致。 结论 成功地构建了结核分支杆菌Ag85B基因真核表达株。

Objective:To construct DNA vaccine based on Ag85B gene of M. tuberculosis.Methods:Ag85B gene was amplified by PCR and digested by restriction endonuclease NheⅠ , BamHⅠ , and then inserted into eukaryotic expression vector pVAX1, transferred into E. coli DH5α. The recombinant plasmid was confirmed by PCR,restriction endonuclease digestion and nucleotide sequencing.Results:Using genomic DNA of M. tuberculosis H37Rv strain as template, 978bp was amplified with PCR. PCR of the recombinant plasmid was positive. After the restriction endonuclease digestion, 978bp fragments of Ag85B DNA were visible. Nucleotide sequence identified that the sequence of recombinant plasmid accorded with Ag85B gene reported. Conclusions:M. tuberculosis Ag85B DNA vaccine was successfully constructed. It would be the foundation of further studying prevention and treatment of tuberculosis.