表达鸡TRAIL基因的重组5型腺病毒构建

Construction of recombinant type 5 adenovirus expressing chicken TRAIL gene

本研究采用RT-PCR从鸡脾细胞扩增到TRAIL基因,测序验证后,将扩增的TRAIL基因开放阅读框克隆到pShuttle-CMV上,与pAdeasy-1载体在BJ5183细菌内同源重组,获得含有TRAIL基因的5型腺病毒重组质粒,进而将PacⅠ线性化重组质粒转染AD-293细胞,收获重组腺病毒。并对后者进行RT-PCR扩增,获得TRAIL基因片段,经westernblot鉴定显示构建的重组腺病毒可表达TRAIL蛋白,表明TRAIL基因在构建的重组腺病毒rAd5-TRAIL中得到表达。本研究为下一步的动物试验和研究TRAIL基因的抑瘤作用奠定了基础。

To construct recombinant type 5 adenovirus expressing chicken TRAIL gene, the TRAIL cDNA was obtained by RT-PCR from the total RNA of chicken spleen cells and cloned into pShuttle-CMV vector. Recombinant adenovirus vector expressing TRAIL gene was constructed by homologous recombination between the linearized recombinant shuttle vector and pAdeasy-1 in BJ5183 bacteria. The recombinant type 5 adenovirus was obtained by transfecting Pac I digested recombinant adenovirus vector into AD-293 cells. Western blot anlysis showed that the TRAIL gene was successfully expressed in the recombinant adenovirus.