布鲁氏菌M5-90疫苗株virB2基因缺失株的构建及鉴定

Construction and identification of virB2 deletion mutants of Brucella vaccine strain M5-90

【目的】构建布鲁氏菌M5-90疫苗株virB2基因缺失株。【方法】利用常规分子生物学技术构建自杀载体pGEM-7zf-ΔvirB2-sacB,通过同源重组的方法,将电转化后的布鲁氏菌分别经100 mg/L氨苄抗性筛选和5%蔗糖敏感性筛选,获得基因缺失株。对获得的基因缺失株进行PCR鉴定和稳定性检测。【结果】成功构建M5-90ΔvirB2基因缺失株,并且该缺失株在10代以内未发生回复突变。【结论】为研发新型布鲁氏菌弱毒基因缺失活苗奠定基础。

[Objective]To construct Brucella vaccine strain M5-90 ΔvirB2 mutants.[Methods]Suicide plasmid pGEM-7zf-ΔvirB2-sacB was constructed by traditional molecular biology technology.Through the method of homologous recombination,we screened Brucella gene deletion mutant by 100 mg /L ampicillin resistance screening and 5% sugar sensitivity screening after electroporation.The M5-90 ΔvirB2 mutant was identified by PCR.Its stability was detected by continuous bacteria culture.[Results ] The virB2 deletion mutant strain was successfully constructed.The reverse mutation did not occur within 10 passages.[Conclusion]The results of this study will be based on developing the new attenuated vaccine of Brucella gene deletion mutants.