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不同病毒启动子在指导哺乳动物细胞基因表达中的效率:3'非翻译序列的影响

Efficiency of different viral promoters in directing gene expression in mammalian cells: effect of 3'-untranslated sequences.

作者信息

Rotondaro L, Mele A, Rovera G

机构信息

Department of Biotechnology, Menarini Ricerche Sud, Pomezia (Roma), Italy.

出版信息

Gene. 1996 Feb 12;168(2):195-8. doi: 10.1016/0378-1119(95)00767-9.

Abstract

We compared (i) the enhancer/promoter (mCMV promoter) from the murine cytomegalovirus (CMV) major immediate early gene,(ii) the enhancer/promoter from human CMV major immediate early gene, containing a short promoter (h1CMV) or a long stretch of 5' untranslated region (UTR) from the gene promoter (h2CMV) and (iii) the simian virus 40 (SV40) enhancer/early region promoter (SV2) for their ability to direct foreign gene expression in transiently transfected mammalian cell lines. Two series of recombinant plasmids containing the different viral promoters fused to the cat reporter gene and 3'-UTR for processing of transcripts from either the SV40 early region or the rabbit Beta 1-globin-encoding gene (Glb) were also analyzed for their effect on transient gene expression. The mCMV was the most active in dihydrofolate reductase-deficient Chinese hamster ovary (CHOdhfr-) cells and BALB/3T3 clone A31 mouse embryo cells. The h2CMV was more active than the other promoters in Bowes human melanoma cells and in Vero African green monkey kidney cells. In human hepatoma (Hep G2) cells, similar levels of CAT synthesis were observed with the h2CMV- and the mCMV-based vectors. In Hep G2 and Bowes cells, 3'-UTR from the SV40 early region resulted in consistently higher levels of cat expression, as compared to the rabbit beta 1-Glb gene, while the converse was true in BALB/3T3 clone A31 and Vero cells. SV40 early region and rabbit beta1-Glb gene 3'-UTR resulted in similar cat expression in CHOdhfr- cells.

摘要

我们比较了

(i) 来自鼠巨细胞病毒 (CMV) 主要立即早期基因的增强子/启动子(mCMV 启动子);(ii) 来自人 CMV 主要立即早期基因的增强子/启动子,其包含一个短启动子(h1CMV)或来自该基因启动子的一段长的 5' 非翻译区(UTR)(h2CMV);以及 (iii) 猿猴病毒 40(SV40)增强子/早期区域启动子(SV2)在瞬时转染的哺乳动物细胞系中指导外源基因表达的能力。还分析了两系列重组质粒,其包含与氯霉素乙酰转移酶(cat)报告基因融合的不同病毒启动子以及用于处理来自 SV40 早期区域或兔β1 - 珠蛋白编码基因(Glb)转录本的 3' - UTR,以研究它们对瞬时基因表达的影响。mCMV 在二氢叶酸还原酶缺陷的中国仓鼠卵巢(CHOdhfr -)细胞和 BALB/3T3 克隆 A31 小鼠胚胎细胞中活性最高。h2CMV 在鲍氏人黑色素瘤细胞和非洲绿猴肾 Vero 细胞中比其他启动子更具活性。在人肝癌(Hep G2)细胞中,基于 h2CMV 和 mCMV 的载体观察到相似水平的 CAT 合成。在 Hep G2 和鲍氏细胞中,与兔β1 - Glb 基因相比,来自 SV40 早期区域的 3' - UTR 导致 cat 表达水平持续更高,而在 BALB/3T3 克隆 A31 和 Vero 细胞中情况则相反。SV40 早期区域和兔β1 - Glb 基因 3' - UTR 在 CHOdhfr - 细胞中导致相似的 cat 表达。

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