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目的 :对鼠疫耶尔森氏菌F1抗原在鼠伤寒沙门氏菌中的表达及其保护力进行研究。方法 :PCR、化学转化及电穿孔法。结果 :采用PCR方法 ,特异地扩增了鼠疫菌F1抗原caf操纵子的 5Kb基因片段。将PCR产物与质粒载体连接后 ,经化学方法转化大肠杆菌LE392 ,获得caf基因克隆子 ,所得克隆子能较好地表达F1抗原基因。应用电穿孔法将克隆子的重组质粒转入鼠伤寒沙门氏菌G30内获得转化子 ,其F1抗原表达水平与在大肠杆菌LE392中的表达水平相同 ,反向血凝滴度可达 1∶80以上 ,与EV株相近。用该株免疫小鼠 ,2 0d后用 141株攻击 ,可延长平均死亡时间。结论 :caf基因的克隆子在鼠伤寒沙门氏菌G30中能够有效表达 ,且对小鼠显示出保护效果。该研究为构建新型鼠疫菌苗提供了线索
Objective: To study the expression of Yersinia pestis F1 antigen in Salmonella typhimurium and its protective ability. Methods: PCR, chemical transformation and electroporation. Results: The 5Kb gene fragment of caf operon of Y. pestis F1 antigen was amplified by PCR. After the PCR product was ligated with the plasmid vector, the E. coli LE392 was chemically transformed to obtain the caf gene clones, and the resulting clones could express the F1 antigen gene better. The recombinant plasmid was transformed into Salmonella typhimurium G30 by electroporation to obtain the transformant. The expression level of F1 antigen was the same as that in E. coli LE392, and the reverse titer of the clot was up to 1:80. Similar to EV strains. Mice were immunized with this strain and challenged with 141 strains after 20 days to extend the mean time to death. CONCLUSION: The cloned caf gene is expressed efficiently in Salmonella typhimurium G30 and shows protective effect on mice. This study provides a clue to construct a new type of plague vaccine