Assessment of a modular transfection system based upon cellular localization of DNA
Assessment of a modular transfection system based upon cellular localization of DNA
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DOI：
10.1021/mp049969a
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发表时间：
2004-07-01
影响因子：
4.9
通讯作者：
Genileinhart, Richard A.
中科院分区：
文献类型：
--
作者：
Guoi, Chunqiang;Genileinhart, Richard A.
在哺乳动物细胞中递送用于蛋白质生产的质粒DNA不是一个高效的过程。在这项研究中，对质粒DNA的细胞定位可提高转染效率这一理论进行了检验，重点是了解三元转染复合物各成分在细胞中的结合情况。哺乳动物细胞主要从其局部环境摄取转染试剂 - DNA复合物。通过形成由DNA - 转染试剂复合物和重粒子（如二氧化硅）组成的三元复合物，转染效率大幅提高。我们对用三元复合物转染的细胞进行了分析，以确定仅仅沉降是否会影响包含复合物或复合物特定成分的细胞百分比。很大一部分细胞与三元复合物结合，包括二氧化硅纳米颗粒。使用三元复合物对与DNA结合的细胞百分比没有显著影响。这一结果表明，二氧化硅纳米颗粒不仅仅是一种沉降剂，还是一种辅助转染试剂。这些数据还证实，细胞可能包含转染试剂 - DNA复合物，但不表达目的蛋白。这一知识将用于进一步的研究，以更好地设计转染试剂，提高蛋白质生产效率。
Delivery of plasmid DNA for protein production in mammalian cells is not an efficient process. In this study, the theory that cellular localization of plasmid DNA increases transfection efficiency is examined with an emphasis on the understanding of the cellular association of the components of a ternary transfection complex. Mammalian cells take up transfection reagent-DNA complexes primarily from their local environment. Via formation of a ternary complex consisting of the DNA-transfection reagent complex and a heavy particle, such as silica, the efficiency of transfection is substantially increased. We have analyzed cells transfected with the ternary complexes to determine if sedimentation alone affects the percentage of cells that contain the complexes or specific components of the complex. A significant fraction of cells associate with the ternary complexes, including silica nanoparticles. The percentage of cells that associate with DNA was not significantly influenced by the use of the ternary complex. This result suggests that the silica nanoparticles are more than just a sedimentation agent, being also a secondary transfection reagent. These data also confirm that cells may contain transfection reagent-DNA complex but do not express the protein of interest. This knowledge will be used in further research to better design transfection reagents that will increase the efficiency of protein production.