摘要
为了构建布鲁氏菌16MΔomp31基因缺失株,采用PCR方法分别从亲本株16 M上扩增omp31基因的侧翼看序列及枯草芽孢杆菌SacB基因,并将所得片段与pMD18-T载体相连并测序,利用双酶切的方法分别将其连入自杀载体pGEM-7zf+,获得亚克隆pGEM-7zf+-Δomp31-SacB。将所构建好的自杀载体通过电转化入布鲁氏菌16M感受态细胞中,经2次同源重组后筛选出16MΔomp31基因缺失株,并对获得缺失株进行遗传稳定性检测。结果显示:成功获得布鲁氏菌16MΔomp31基因缺失株,该缺失株在15代内未发生回复性突变。本研究为今后研究布鲁氏菌抗凋亡机制奠定基础。
To construct the omp31 deletion mutant of Brucella melitensis 16M,the upstream and downstream of the omp31 gene and SacB gene were amplified by PCR from Brucella melitensis 16M and Bacillus subtilis.After constructing omp31-SacB recombinant fragment into plasmid 18-T simple vector,the suicide plasmid pGEM-7zf＋-Δomp31-SacB was further obtained and transformed into Brucella melitensis 16M by electroporation.The Δomp31 mutant strain was screened out by homologous recombination and its stability was detected by continuous bacteria culture.The results showed that Brucella melitensis 16M Δomp31 mutant strain was successfully generated and reversion was not observed in 15 generations.This research lays a foundation of further study on the anti-apoptosis mechanism and construction of new types of vaccines of Brucella.
出处
《石河子大学学报（自然科学版）》
CAS
2013年第1期30-34,共5页
Journal of Shihezi University(Natural Science)
基金
国家重点基础理论研究发展计划(973计划)项目(2010CB530203)
国家自然科学基金项目(30800813
31060334)
新疆兵团博士基金项目(2009JC15)
石河子大学高层次人才科研启动项目(RCZX200828)
