摘要
目的 构建和鉴定日本血吸虫重组两歧双歧杆菌（Bifidobacterium bifidum,Bb）pGEX-Sj32疫苗.方法 从重庆医科大学附属第一医院传染病寄生虫病研究所构建的重组大肠埃希菌BL21 （pET28α-Sj32）中抽提质粒pET28α-Sj32,通过PCR扩增Sj32抗原编码基因;将Sj32基因定向克隆至大肠埃希菌-双歧杆菌穿梭表达载体pGEX-1λT中,构建重组质粒pGEX-Sj32,将该重组质粒转化至大肠埃希菌BL21感受态细胞（DE3）,抽提质粒后进行双酶切鉴定;采用电穿孔法,将重组质粒pGEX-Sj32转化至Bb,构建Bb（pGEX-Sj32）疫苗,再从该疫苗中抽提重组质粒pGEX-Sj32,以其为模板进行PCR鉴定.结果 从抽提质粒pET28α-Sj32中PCR扩增出长度约为1 270 bp的Sj32基因片段;重组质粒pGEX-Sj32经双酶切鉴定,获得长度为4 947 bp的载体片段和长度为1 270 bp的Sj32基因片段;以抽提的疫苗质粒pGEX-Sj32为模板进行PCR扩增,得到了长度约为1 270 bp的Sj32基因片段,与预期结果相符.结论 成功构建了日本血吸虫重组Bb（pGEX-Sj32）疫苗.
Objective To construct and identify recombinant vaccine Bifidobacterium bifidum （pGEX-Sj32） of Schistosomajaponicum （Sj）.Methods The Sj32 gene amplified by PCR from template of plasmid pET28α-Sj32 that extracted from recombinant Escherichia coli（E.coli） BL21 （pET28α-Sj32） stored in our laboratory was cloned into E.coli-Bifidobacterium bifidum shuttle expression vector pGEX-1λT to construct recombinant plasmid pGEX-Sj32.The recombinant plasmid was transformed into E.coli BL21 （DE3）.The plasmid was extracted and identified by restriction enzyme digestion.The recombinant Bifibacterium bifidum （pGEX-Sj32） vaccine was constructed by electroporating pGEX-Sj32 into Bb.The extracted plasmid was amplified and identified by PCR.Results The gene Sj32 of 1 270 bp in length was amplified by PCR.The restriction endonuclease digestion showed that the length of plasmid vector was 4 947 bp,and Sj32 gene was 1 270 bp.The Sj32 gene amplified by PCR from the template of pGEX-Sj32 extracted from recombinant Bb （pGEX-Sj32） vaccine was 1 270 bp in length which consistent with expected result.Conclusion The recombinant Bb （pGEX-Sj32） vaccine of Sj is successfully constructed.
出处
《中华地方病学杂志》
CAS
CSCD
北大核心
2015年第1期37-40,共4页
Chinese Journal of Endemiology
基金
重庆市科委地方病重大专项基金（2008AB5055、2008AB5008、2008AB5054）
关键词
血吸虫
日本
重组
遗传
两歧双歧杆菌
疫苗
Schistosoma japonicum
Recombination,genetic
Bifidobacterium bifidum
Vaccines
