[1]罗丽芸,白敏,张剑,等.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[J].南京师范大学学报(自然科学版),2015,38(03):64.
 LuoLiyun,BaiMin,ZhangJian,et al.ConstructionofRecombinantAdenovirusofhAIF-1GeneandItsEffectonProliferationandMigrationofA549Cells[J].Journal of Nanjing Normal University(Natural Science Edition),2015,38(03):64.
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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()
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《南京师范大学学报》(自然科学版)[ISSN:1001-4616/CN:32-1239/N]

卷:
第38卷
期数:
2015年03期
页码:
64
栏目:
生命科学
出版日期:
2015-09-30

文章信息/Info

Title:
ConstructionofRecombinantAdenovirusofhAIF-1GeneandItsEffectonProliferationandMigrationofA549Cells
作者:
罗丽芸白敏张剑曹令森曹祥荣
南京师范大学生命科学学院,江苏省分子医学生物技术重点实验室,江苏南京210023
Author(s):
LuoLiyunBaiMinZhangJianCaoLingsenCaoXiangrong
SchoolofLifeSciences,NanjingNormalUniversity,JiangsuProvinceKeyLaboratoryforMolecularandMedicalBiotechnology,Nanjing210023,China
关键词:
hAIF-1腺病毒增殖迁移
Keywords:
hAIF-1adenovirusproliferationmigration
分类号:
Q28
文献标志码:
A
摘要:
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.
Abstract:
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.

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备注/Memo

备注/Memo:
收稿日期:2014-12-04.
基金项目:.企业委托项目(K11100BY62).
通讯联系人:曹祥荣,教授,博导,研究方向:肿瘤发生与基因治疗分子机制.E-mail:08085@njnu.edu.cn
更新日期/Last Update: 2015-09-30