摘要
为研制更加安全有效且遗传背景清晰的减毒鼠伤寒沙门菌活疫苗候选株,在单缺失株SL1344Δcrp的基础上,运用自杀性质粒介导的细菌同源重组技术敲除sipB基因,构建了双缺失株SL1344ΔcrpΔsipB。PCR及测序结果表明SL1344ΔcrpΔsipB构建成功。生物学特性研究表明,SL1344ΔcrpΔsipB血清型未发生变化,遗传性稳定。与亲本株SL1344相比,双缺失株的生长速度明显减慢,毒力降低了约99.9%以上。免疫保护效力试验显示,6周龄昆明小鼠口服免疫SL1344ΔcrpΔsipB后第17天,利用鼠伤寒沙门菌野生株攻毒,保护率为60%;免疫后第21天血清抗体效价达到峰值。上述结果为进一步研制安全有效的鼠伤寒沙门菌疫苗弱毒株奠定了基础。
To develop a more efficient and safer live attenuated Salmonella typhimurium vaccine candidate strain,firstly,the recombinant suicide plasmid pREΔsipBlack of 585 bp sipB was constructed.Next,the SL1344ΔcrpΔsipB mutant was selected by two-step method after the recombinant suicide plasmid was conjugated with SL1344Δcrp.The results of PCR and sequencing showed that SL1344ΔcrpΔsipB mutant was successfully constructed.The serotype of SL1344ΔcrpΔsipB mutant was consistent with SL1344Δcrp mutant and SL1344,and the sipB gene was inherited stably.Compared with parent strain SL1344,the growth speed of SL1344ΔcrpΔsipB mutant was more slowly,and the virulence of SL1344ΔcrpΔsipB mutant decreased by 99.9%.The protection rate of SL1344ΔcrpΔsipB mutant was60%in the 6-week-old Kunming mice challenged with S.typhimuriumfield strain on day 17post-immunization.On day 21post-immunization,level of the antibody in serum of the immunized mice peaked.This result laid the foundation for the further development of safe and effective attenuated S.typhimurium vaccines.
出处
《中国兽医科学》
CAS
CSCD
北大核心
2015年第1期61-67,共7页
Chinese Veterinary Science
基金
河南省科技攻关项目(112102110019)
河南科技大学博士启动基金项目(09001580)
河南科技大学青年科学基金项目(C1802)
