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梁硕, 郭彩霞, 王志成, 李艳博, 张海霞, 王剑峰, 龚守良. 辐射诱导Smac表达腺病毒穿梭载体构建及鉴定[J]. 中国公共卫生, 2009, 25(11): 1333-1334. DOI: 10.11847/zgggws2009-25-11-30
引用本文: 梁硕, 郭彩霞, 王志成, 李艳博, 张海霞, 王剑峰, 龚守良. 辐射诱导Smac表达腺病毒穿梭载体构建及鉴定[J]. 中国公共卫生, 2009, 25(11): 1333-1334. DOI: 10.11847/zgggws2009-25-11-30
LIANG Shuo, GUO Cai-xia, WANG Zhi-cheng, . Construction and identification of adenoviral shuttle vector with radiation-inducible smac gene[J]. Chinese Journal of Public Health, 2009, 25(11): 1333-1334. DOI: 10.11847/zgggws2009-25-11-30
Citation: LIANG Shuo, GUO Cai-xia, WANG Zhi-cheng, . Construction and identification of adenoviral shuttle vector with radiation-inducible smac gene[J]. Chinese Journal of Public Health, 2009, 25(11): 1333-1334. DOI: 10.11847/zgggws2009-25-11-30

辐射诱导Smac表达腺病毒穿梭载体构建及鉴定

Construction and identification of adenoviral shuttle vector with radiation-inducible smac gene

  • 摘要: 目的 构建辐射敏感启动子Egr-1诱导人线粒体促凋亡基因(Smac)表达的腺病毒穿梭载体pshuttle-Egr1-hSmac。方法 利用RT-PCR方法从5月龄流产的胎儿肝脏组织中扩增出人Smac DNA编码序列(CDs),从pMD19T-Egr1质粒上将Egr-1基因切下,利用基因重组技术构建含有辐射诱导启动子Egr1的腺病毒穿梭载体pshuttle-Egr1-hSmac。结果 经测序证实,获得的人Smac基因与GenBank公布的完全一致,表明克隆人Smac基因成功;pshut-tle-Egr1-hSmac分别用EcoRⅠ、SmaⅠ和BamHⅠ进行单酶切,片段大小分别为2621和5137;1437,2282和4039;3303和4455 bp,酶切鉴定结果与预期结果完全一致,说明质粒pshuttle-Egr1-hSmac构建正确。结论 成功地克隆了人Smac基因和构建了辐射诱导表达的腺病毒穿梭载体pshuttle-Egr1-hSmac。

     

    Abstract: Objective To construct the adenoviml shuttle vector,the pshuttle-Egrl-hSmac containing radation-sensitive Egrl promoter,and the second mitochondria-derived activator of caspases (Smac) gene.Methods The Smac gene was amplified by reverse transcriptase-polymemse chain reaction (RT-PCR) from fetal liver,and the Egrl promoterwas cut down from plasmid pMD19T-Egrl.Then the adenovirus shuttle vector pshuttle-Egrl-hSmac was constmcted by the gene recombinant technique.Results The sequence of amplified Smac genewas in concordancewith thatpublished on GenBank,indicating that the Smac gene was cloned successfully.Furtheanore,the recombinantplasmidpshuttle-Egrl-Smacwas identified by endonuclease digestion,which could be digested into two or three fragements of EcoRⅠ,SmaⅠ and BamHⅠ,respectively The length of them were 2621 and 5137 base pairs for EcoRⅠ,1437,2282 and 4039 base pairs for SmaⅠ,3303 and 4455 base pairs for BamHⅠ.The results of the identification confirmed that the recombinant plasmid pshuttle-Egrl-Smac was constructed successfully.Conclusion The human Smac gene was cloned and the recom binant plasmid pshuttle-Egrl-Smac was constructed successfully

     

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