FGF21腺病毒表达载体的构建及鉴定  

Construction and identification of FGF21 adenovirus expression vector

作　　者：严莉莉[1] 项丽[1] 庞实锋[1] 段杏华 曹定国[1] 梁朋[1] 张国平[1] 丁银润[1] 胡传银[1] 郑克勤[1] 

机构地区：[1]广东医学院生物学教研室,广东东莞523808 [2]广东省惠阳三和医院,广东惠州516213

出　　处：《广东医学院学报》2015年第1期39-44,共6页Journal of Guangdong Medical College

基　　金：广东省自然科学基金项目(No.S2012040006272)

摘　　要：目的构建成纤维生长因子21(FGF21)表达载体。方法将FGF21基因克隆到腺病毒穿梭质粒p AdTrack经DH5α感受态菌扩增,Pme I酶切后热激转化到含腺病毒骨架质粒p Ad Easy-1的BJ5183感受态菌内同源重组,经卡那霉素筛选及Pac I酶切鉴定,并在DH5α菌中扩增得到Ad-FGF21重组腺病毒质粒,再经Pac I酶切线性化后转染到293A细胞包装成重组腺病毒。结果基因测序结果表明成功合成穿梭载体p Ad-Track-FGF21。PCR和酶切鉴定结果证实含FGF21基因的重组腺病毒表达载体构建成功。Ad-FGF21转染293A细胞并制备出高滴度的重组腺病毒。结论利用腺病毒表达载体获得了携带FGF21的重组腺病毒表达载体。Objective To construct the fibroblast growth factor 12（FGF21） expression vector. Methods FGF21 gene was cloned into shuttle plasmid p Ad-Track and then amplified in DH5 alpha bacteria. After restriction digestion with Pme I, it was transfected into the adenoviral backbone plasmid p Ad Easy-1 in BJ5183 competent bacteria by heat method. Ad-FGF21 was selected by kanamycin and identified by Pac I restriction enzyme, followed by amplified in DH5 alpha bacteria. Ad-FGF21 plasmid was linearized by Pac I and then transfected into 293 A cells. Results The shuttle vector p Ad-Track-FGF21 and recombinant adenovirus FGF21 expression vector were successfully constructed by gene sequencing, PCR and restriction enzyme digestion, respectively. The high-titer recombinant adenovirus was prepared by transfection with 293 A cells. ConclusionRecombinant adenovirus FGF21 expression vector can be successfully constructed using adenovirus vector system.

关 键 词：腺病毒 成纤维细胞生长因子 基因治疗 

分 类 号：R587.1[医药卫生—内分泌]