表达mIFN-γ的重组链球菌的构建及其对重组Sj-F1链球菌疫苗免疫调节作用研究  

Construction of recombinant streptococcus expressing mIFN-γ and immunoregulation on recombinant Sj-F1 streptococcus

作　　者：彭丹[1] 杨明[1] 刘玮[1] 陈利玉[1] 

机构地区：[1]中南大学湘雅医学院微生物学系,湖南长沙410078

出　　处：《中国现代医学杂志》2013年第9期28-33,共6页China Journal of Modern Medicine

基　　金：湖南省自然科学基金重点项目(No:10JJ2012);湖南省研究生科研创新项目(No:2011ssxt103)

摘　　要：目的构建表达mIFN-γ的重组链球菌,并与重组日本血吸虫Sj-F1链球菌疫苗进行联合免疫,观察IFN-γ对重组日本血吸虫Sj-F1链球菌疫苗的免疫调节效应。方法用PCR法扩增mIFN-γ基因,亚克隆至链球菌重组质粒pGMB1,经PCR、限制性酶切和测序鉴定筛选阳性重组子。将阳性重组质粒转化链球菌载体,经抗生素抗性转化及染色体PCR鉴定筛选阳性重组链球菌。用Streak blot和Western blot分析鉴定重组链球菌的表达情况。鉴定后的重组mIFN-γ链球菌与重组Sj-F1链球菌疫苗联合应用,经滴鼻接种免疫昆明小鼠,观察诱导产生的减虫和减卵效果。结果成功构建了分泌mIFN-γ重组链球菌。Streak blot和Western blot分析表明重组链球菌能分泌mIFN-γ并可稳定遗传。免疫保护性实验表明,重组mIFN-γ链球菌与重组日本血吸虫Sj-F1链球菌疫苗联合免疫,减虫率和减卵率分别为31.56%和47.48%,与单用重组Sj-F1链球菌疫苗比较减虫率和减卵率圴增加。结论结果表明,重组mIFN-γ链球菌能增强重组Sj-F1链球菌疫苗抗日本血吸虫感染的保护力。【Objective】 To construct a recombinant Streptococcus Expressing mIFN-γ and study its immunoregulation on recombinant Schistosoma japonicum Sj-F1 Streptococcus vaccine against challenge infection in mice with Schistosom japonicum.【Motheds】 The mIFN-γ gene was amplified by PCR and subcloned into Streptococcal plasmid vector pGMB1.The positive recombinant was identified by PCR,restriction enzyme digestion and sequencing.Then the recombinant plasmid was transformed into Streptococcal recipient.The recombinant Streptococcus was screened with antibiotics and PCR.The mIFN-γ secreted by the recombinant Streptococcus was identified with Streak blot and Western blot.Recombinant Streptococcus expressing mIFN-γ was used to immunize Kunming mice intranasally together with recombinant Sj-F1Streptococcus vaccine.Two weeks after the third vaccination mice were challenged with S.japonicum cercariae.The immunoprotection was assessed by worm and egg reduction percentage.【Results】 The recombinant mIFN-γ Streptococcus was successfully constructed.Immunization experiment showed co-immunization with recombinant mIFN-γ Streptococcus and recombinant Sj-F1 Streptococcus provided 31.56% worm reduction rates and 47.48% egg reduction rates in mice.Compared with that of mice administrated with only recombinant Sj-F1 Streptococcus,the worm and egg reduction rates in co-immunized mice were increased.【Conclusion】 The recombinant mIFN-γ Streptococccus can express mIFN-γ.The recombinant mIFN-γ Streptococcus can significantly enhance the protective immunity against S.japonicum infection induced by recombinant Sj-F1 Streptococcus vaccine.

关 键 词：日本血吸虫 mIFN-γ Sj-F1 重组链球菌 免疫 

分 类 号：R373[医药卫生—病原生物学]