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  • Title: Insertion of nuclear factor-kappaB binding sequence into plasmid DNA for increased transgene expression in colon carcinoma cells.
    Author: Thanaketpaisarn O, Nishikawa M, Okabe T, Yamashita F, Hashida M.
    Journal: J Biotechnol; 2008 Jan 01; 133(1):36-41. PubMed ID: 17961782.
    Abstract:
    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.
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