携带小鼠CD40Ig基因重组腺病毒的构建及鉴定

Construction and identification of adenovirus vector containing mouse CD40Ig gene

目的构建携带小鼠CD4 0Ig基因的重组腺病毒载体 ,为研究供体特异性移植耐受诱导做准备。方法分别克隆小鼠CD4 0胞外区cDNA和IgG2a Fc段cDNA ,以PCR方法将二者连接为CD4 0Ig。再将后者装入腺病毒穿梭质粒 pAdTrack CMV ,与骨架质粒在大肠杆菌BJ5 183中同源重组 ,经 2 93细胞包装、扩增后得到携带CD4 0Ig的重组腺病毒AdCD4 0Ig。 结果成功构建携带CD4 0Ig的重组腺病毒载体系统 ,病毒滴度为 5× 10 9efu/ml。结论构建的AdCD4 0Ig可用于供体特异性移植耐受的实验研究。

Objective To construct recombinant adenovirus vector containing mouse CD40Ig fusion gene for the study of induction of donor-specific tolerance. Methods CD40Ig fusion gene was constructed by PCR overlapping technique, and was cloned into the shuttle plasmid pAdTtrack-CMV. The linearized shuttle plasmid was co-transfected into the E.coli strain BJ5183 with bone plasmid pAdeasy1, then the recombinant adenovirus plasmid was generated. The adenovirus was packaged and amplified in Cells 293. Results The recombinant virus AdCD40Ig was successfully constructed and its titer was 5×109 efu/ml. Conclusion AdCD40Ig can be used as an agent in experiment to induce donor-specific tolerance.