文章编号：2095-1191（2019）06-1356-06
Abstract：【Objective】The aim of the study was to construct an adenoviral vector containing the telomerase catalytic subunit（hTERT） gene， optimize its packaging conditions and lay a foundation for further study of the interaction mechanism between telomerase and cell reprogramming. 【Method】The adenoviral vector containing the hTERT gene was constructed by using the AdEasy-1 system of recombinant adenovirus， and then it was packaged in HKE293A cells by liposome， electroporation and different serum concentrations of fetal bovine. The optimal packaging condition was selected. Recombinant adenovirus rAd-hTERT was collected and infected mouse fetal fibroblasts in vitro， and activity function of recombinant adenovirus rAd-hTERT was verified. 【Result】The recombinant adenoviral vector（pAd-hTERT） carrying hTERT gene was successfully constructed with recombinant adenovirus rAd-hTERT as skeleton plasmid. The optimal packaging condition was liposome method combining with 15% serum of fetal bovine. The titer of virus obtained under such condition was the highest（1×1011 IFU/mL）. After 72 h of adenovirus（rAd-hTERT） carrying hTERT gene infected mouse fetal fibroblasts， target gene bands of 3450 bp was amplified， which meaned that hTERT gene was successfully expressed in mouse fetal fibroblasts. 【Conclusion】The recombinant adenovirus carrying hTERT gene has been successfully prepared by liposome method combining with 15% serum of fetal bovine. It has high virus titer. Infecting mouse fetal fibroblasts can successfully express hTERT gene， which further verifies that telomerase has biological activity of functional protein.
Key words： telomerase; adenovirus; cell reprogramming; vector construction; virus packaging; functional verification
收稿日期：2019-04-04
基金项目：国家自然科学基金项目（31560331，31660340）;内蒙古自然科学基金项目[2017MS（LH）0213，2017MS0337]
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