%0 Journal Article %T hAIF-1基因重组腺病毒的构建及其对A549细胞增殖和迁移的影响 %A 罗丽芸 %A 白敏 %A 张剑 %A 曹令森 %A 曹祥荣 %J 南京师范大学学报(自然科学版) %D 2015 %X hAIF-1是一种炎症因子,本研究通过构建hAIF-1基因的重组腺病毒,探讨hAIF-1对A549细胞增殖和迁移能力的影响.用PCR的方法克隆hAIF-1基因编码序列,并亚克隆到腺病毒穿梭质粒载体pDC315-EGFP上,构建穿梭质粒pDC315-EGFP-hAIF-1,将穿梭质粒与腺病毒骨架质粒pBHGloxdeltaE13Cre共转染293细胞,利用AdMax腺病毒载体包装系统包装腺病毒,TCID50法测定病毒滴度,利用RT-PCR和WesternBlot分析检测目的基因表达,用MTT法检测A549细胞的增殖,细胞划痕和Transwell实验检测A549细胞的迁移.结果表明,成功构建了表达hAIF-1基因的重组腺病毒,所得病毒滴度为2.5×108pfu/mL;用hAIF-1重组腺病毒感染人A549细胞,RT-PCR和WesternBlot鉴定显示细胞可过表达hAIF-1;MTT显示过表达hAIF-1可显著增强A549细胞增殖能力;细胞划痕和Tranwell实验证明hAIF-1能够促进A549细胞迁移.</br>hAIF-1isaninflammatoryfactor.ThestudywassupposedtoexploretheeffectofhAIF-1ontheproliferationandmigrationofA549cellsbythepreparationoftherecombinantadenoviruswithhAIF-1gene.ThehAIF-1genewasclonedbyPCRandsubclonedintotheshuttlevectorpDC315-EGFP.TheshuttleplasmidandadenovirusgenomicplasmidpBHGloxdeltaE13Crewereco-transfectedinto293cellsforpackagingrecombinantadenoviruswiththehelpoftheadenoviruspackagingsystemAdMax.ThetiteroftheviruswasmeasuredbyTCID50.TheexpressionofthetargetgenewasidentifiedbyRT-PCRandWesternBlot,andtheeffectofthetargetgeneontheproliferationandmigrationofA549cellswasexaminedbyMTT,woundhealingandTranswellassay.TheresultsshowedthattheAdDC315-EGFP-hAIF-1wassuccessfullyconstructedandthetiteroftheviruswas2.5×108pfu/mL;AnalysisofRT-PCRandWesternBlotindicatedthatA549cellscouldoverexpresshAIF-1afterinfectedwiththerecombinantadenovirus;TheMTTassayshowedthathAIF-1couldincreasetheproliferativerateofA549cells;ThewoundhealingandTranswellassayprovedthathAIF-1couldpromotethemigrationofA549cells %K hAIF-1 %K 腺病毒 %K 增殖 %K 迁移< %K /br> %K hAIF-1 %K adenovirus %K proliferation %K migration %U http://njsfdxzrb.paperonce.org/oa/darticle.aspx?type=view&id=201503008