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首页> 外文期刊>Journal of Biotechnology >Insertion of nuclear factor-kappaB binding sequence into plasmid DNA for increased transgene expression in colon carcinoma cells
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Insertion of nuclear factor-kappaB binding sequence into plasmid DNA for increased transgene expression in colon carcinoma cells

机译:将核因子-κB结合序列插入质粒DNA以提高结肠癌细胞中的转基因表达

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摘要

To increase plasmid DNA (pDNA)-based transgene expression, 5, 10 or 20 repeats of nuclear factor kappaB (NF-kappaB) binding sequences were inserted upstream of the cytomegalovirus (CMV) promoter region of a conventional pDNA encoding firefly luciferase (pCMV-Luc) to obtain pCMV-kappaB5-Luc, pCMV-kappaB10-Luc and pCMV-kappaB20-Luc. Murine carcinoma colon 26 cells, in which NF-kappaB was constitutively activated, were co-transfected with a firefly luciferase-expressing pDNA and a renilla luciferase-expressing pDNA having no NF-kappaB binding sequences using cationic liposomes. The expression efficiency of pCMV-kappaB(n)-Luc was evaluated using the ratio of the luciferase activities. Increasing numbers of NF-kappaB binding sequences significantly increased transgene expression. The expression was increased by NF-kappaB activators and the effects were marked with pDNA having many NF-kappaB binding sequences. These results indicate that insertion of NF-kappaB binding sequences into pDNA is an effective approach to increase transgene expression in cancer cells in which NF-kappaB is activated.
机译:为了增加基于质粒DNA(pDNA)的转基因表达,将5、10或20个重复的核因子kappaB(NF-kappaB)结合序列插入编码萤火虫荧光素酶(pCMV- Luc)获得pCMV-κB5-Luc,pCMV-κB10-Luc和pCMV-κB20-Luc。使用阳离子脂质体将其中NF-kappaB被组成性激活的鼠癌结肠26细胞与表达萤火虫荧光素酶的pDNA和不具有NF-kappaB结合序列的表达雷尼尔荧光素酶的pDNA共转染。使用荧光素酶活性的比率评估pCMV-κB(n)-Luc的表达效率。 NF-κB结合序列数目的增加显着增加了转基因表达。 NF-kappaB激活剂增加了表达,并用具有许多NF-kappaB结合序列的pDNA进行了标记。这些结果表明,将NF-κB结合序列插入pDNA是增加其中NF-κB被活化的癌细胞中转基因表达的有效方法。

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