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Construction of a survivin promoter-mediated adeno-associated virus expression vector carrying a fusion gene constituted with cytosine deaminase and survivin mutant

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Author:
No author available
Journal Title:
WORLD CHINESE JOURNAL OF DIGESTOLOGY
Issue:
11
DOI:
10.3969/j.issn.1009-3079.2009.11.005
Key Word:
Survivin启动子;Survivin突变体;胞嘧啶脱氨酶基因;载体构建

Abstract´╝Ü 目的:构建Survivin启动子介导自杀基因胞嘧啶脱氨酶(CD)与Survivin Cys84Ala突变体(SurMut)融合基因的重组腺相关病毒质粒(pAM/SurP-CDglySurMut),探讨其在肿瘤细胞中有效表达融合基因的能力.方法:PCR扩增E.coli CD基因和带有甘氨酸(gly)连接体的Survivin突变体片段,亚克隆至具有Survivin启动子的腺相关病毒载体pAM/SurP,获得重组表达载体pAM/SurP-CDglySurMut.重组载体瞬时转染胃癌细胞,Western blot检测融合蛋白的表达.结果:经PCR扩增成功克隆E.coli CD基因和带有gly连接体的Survivin突变体片段.重组阳性克隆经限制性酶切鉴定和基因测序,证实目的基因CDglySurMut已正确插入pAM/SurP载体中.Western blot证实Survivin启动子能够驱动CDglySurMut融合基因在胃癌细胞中达.结论:pAM/SurP-CDglySurMut载体能够在肿瘤细胞中表达融合蛋白,为进一步探讨其靶向治疗肿瘤奠定了实验基础

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