小鼠SIGIRR基因重组腺病毒的构建及鉴定  

Construction and Identification of Mouse SIGIRR Gene Recombinant Adenovirus

作　　者：梁晓华[1] 王艳[1] 田丰[1] 黄勋[1] 高坤祥[1] 李小飞[1] 姜涛[1] 

机构地区：[1]第四军医大学唐都医院胸腔外科,陕西西安710038

出　　处：《动物医学进展》2013年第10期1-8,共8页Progress In Veterinary Medicine

基　　金：国家自然科学基金项目(81070062)

摘　　要：拟构建小鼠SIGIRR基因重组腺病毒并进行鉴定,为SIGIRR在小鼠急性肺损伤预防及治疗方面的研究提供基础。利用AdEasyTM系统构建SIGIRR基因重组腺病毒,并检测SIGIRR mRNA及蛋白在感染细胞及组织中的表达。通过多次扩增后,大量制备高滴度腺病毒。结果表明,本试验成功构建了携带小鼠源性SIGIRR基因的重组腺病毒表达载体Ad-mSIGIRR,病毒感染滴度为5×1010 pfu/mL,滴度符合下一步试验要求。Western blot和免疫组化方法分别检测到mSIGIRR在体内的高效表达,且经气管滴注感染小鼠后,不会引起正常小鼠肺组织发生病理变化。明确了SIGIRR基因的重组腺病毒可以在小鼠活体内使mSIGIRR过表达,且经气管滴注5×109pfu的病毒感染小鼠后,不会引起正常小鼠肺组织发生病理变化。The mouse SIGIRR gene recombinant adenovirus was constructed and identified as the basic re- search of SIGIRR in the prevention and treatment of acute lung injury（ALI）. AdEasyTM system was used to construct the mouse SIGIRR gene recombinant adenovirus,and detect the SIGIRR mRNA and SIGIRR protein expression in infected cells and tissues. A high-titer adenovirus was prepared by repeated amplifica- tion. The result showed that mouse SIGIRR gene recombinant adenovirus （Ad-mSIGIRR）was successfully constructed and the titer is 5×10^10 pfu/mL which matched the requirement of next experiment. After intra- tracheal instillation of mouse SIGIRR gene recombinant adenovirus, SIGIRR protein over-expression was found in mouse lung tissue by Western blot and immunohistochemistry, and did not cause pathological le- sions in normal mouse lung tissues with the experimental dose. This study confirmed that mouse SIGIRR gene recombinant adenovirus could enhance the expression of SIGIRR in mice after intratracheal instilla- tion, and did not cause pathological lesions in normal mouse lung tissues with the experimental dose 5 ×10^10 puf/mL.

关 键 词：单免疫球蛋白白细胞介素1受体相关蛋白（SIGIRR） 急性肺损伤 腺病毒载体 AdEasyTM Toll样受体（TLR） 

分 类 号：Q784[生物学—分子生物学]