人白介素24基因重组腺病毒载体的构建与鉴定

目的构建含有人白介素24(hIL-24)基因的重组腺病毒载体,为下一步病理性瘢痕的基因治疗研究奠定实验基础。方法采用基因工程技术将hIL-24基因的cDNA亚克隆至穿梭质粒pAdTrack-CMV上,鉴定正确后在PAdEasy系统中进行细菌内同源重组,通过脂质体将正确重组体包裹并转染293A细胞以包装并扩增病毒。采用酶切及PCR方法对重组腺病毒进行鉴定。结果酶切和PCR结果证实hIL-24基因重组腺病毒载体构建成功并可在293A细胞中表达,病毒滴度达107pfu/ml。结论成功构建了有较强感染能力的含hIL-24基因的重组腺病毒载体。

The construction and Identification of Recombinant Adenovirus Vector of hIL-24

Objective To construct and identify the recombinant adenovirus vector of human IL-24 gene for future gene therapy of pathological scar tissue. Methods The human IL-24 gene fragment was cloned into the shuttle plasmid pAdTrack-CMV to form the transfer vector by the method of homogenous recombination in bacteria of PAdEasy system. Then the right recombinant adenovirus was transfected into 293A cells using Lipofectine 2000. The recombinant adenovirus vector was identified by restriction enzyme digestion and polymerase chain reaction (PCR). Results Restriction endonuclease and PCR analysis confirmed that the human IL-24 gene was successfully inserted into the adenovirus vector. The titer of the recombinant adenovirus was 107pfu/ml. Conclusion The recombinant adenovirus vector of hIL-24 was successfully constructed with highly infection.