Electroporation-mediated gene transfer efficiency is reduced by linear plasmid carrier DNAs.
Electroporation-mediated gene transfer efficiency is reduced by linear plasmid carrier DNAs.
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DOI：
10.1016/0003-2697(92)90429-b
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发表时间：
1992-09
影响因子：
2.9
通讯作者：
J. Nickoloff;R. J. Reynolds
中科院分区：
文献类型：
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作者：
J. Nickoloff;R. J. Reynolds
载体DNA通常被发现通过磷酸钙共沉淀刺激DNA介导的中国仓鼠卵巢（CHO）细胞的基因转移。然而，在采用电穿孔的研究中，我们观察到线性质粒DNA对CHO细胞的转染具有抑制作用。这个意想不到的结果促使我们探索各种类型和形式的质粒、粘粒和染色体DNA对转染效率的影响。载体DNA的形式和类型都影响转染效率。质粒载体DNA的环状和线性形式具有相反的效果：环状质粒增加，线性质粒降低转染效率。这些影响是独立的同源性与选定的质粒，可能是独立的同源重组机制。细菌基因组DNA未能刺激转染，而小牛胸腺和粘粒DNA主要由人类序列刺激转染显着。我们的研究结果具有重要意义的质粒为基础的实验在哺乳动物细胞中，如那些涉及诱导质粒间同源重组。
Carrier DNA has generally been found to stimulate DNA-mediated gene transfer of Chinese hamster ovary (CHO) cells by calcium phosphate coprecipitation. In studies employing electroporation, however, we observed that linear plasmid DNA was inhibitory to the transfection of CHO cells. This unexpected result prompted us to explore the effects of various types and forms of plasmid, cosmid, and chromosomal DNAs on transfection efficiencies. Both carrier DNA form and type were found to influence transfection efficiencies. Circular and linear forms of plasmid carrier DNA had opposite effects: circular plasmids increased and linear plasmids decreased transfection efficiencies. These effects were independent of homology with the selected plasmid and are probably independent of homologous recombination mechanisms. Bacterial genomic DNA failed to stimulate transfection, while calf thymus and cosmid DNA consisting primarily of human sequences stimulated transfection significantly. Our results have importance for plasmid-based experiments in mammalian cells such as those involving the induction of interplasmid homologous recombination.