A rapid and efficient polyethylenimine-based transfection method to prepare lentiviral or retroviral vectors: useful for making iPS cells and transduction of primary cells
A rapid and efficient polyethylenimine-based transfection method to prepare lentiviral or retroviral vectors: useful for making iPS cells and transduction of primary cells
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一种快速有效的基于聚乙烯亚胺的转染方法，用于制备慢病毒或逆转录病毒载体：可用于制备 iPS 细胞和原代细胞转导。
DOI：
10.1007/s10529-016-2123-2
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发表时间：
2016-09-01
影响因子：
2.7
通讯作者：
Sun, Pingnan
中科院分区：
文献类型：
--
作者：
Yang, Shaozhe;Shi, Haijun;Sun, Pingnan
为了提高PEI转染法的效率、重复性和一致性，研究了PEI或DNA浓度、PEI/DNA稀释液、PEI/DNA复合物制备方式、血清影响、PEI/DNA复合物孵育时间和转染时间等因素对转染效率的影响。在PEI/DNA转染复合物的制备过程中温和混合对于高转染效率是至关重要的。PEI可在室温或4 A ℃下储存，最重要的是应避免多重凝胶化。基于PEI的新方法在不同类型细胞中的转染效率，如293 T，Cos-7，HeLa，HepG 2，Hep 3B，Huh 7和L02，也高于先前的方法。优化后的方法生产的慢病毒和逆转录病毒的滴度分别是原方法的10倍和3倍，为制备重组慢病毒和逆转录病毒载体提供了一种快速、有效的方法。
To improve the efficiency, reproducibility and consistency of the PEI-based transfection method that is often used in preparation of recombinant lentiviral or retroviral vectors.The contributions to transfection efficiency of multi-factors including concentration of PEI or DNA, dilution buffer for PEI/DNA, manner to prepare PEI/DNA complexes, influence of serum, incubation time for PEI/DNA complexes, and transfection time were studied. Gentle mixing during the preparation of PEI/DNA transfection complexes is critical for a high transfection efficiency. PEI could be stored at room temperature or 4 A degrees C, and most importantly, multigelation should be avoided. The transfection efficiency of the PEI-based new method in different types of cells, such as 293T, Cos-7, HeLa, HepG2, Hep3B, Huh7 and L02, was also higher than that of the previous method. After optimization, the titer of our lentiviral system or retroviral system produced by PEI-based new method was about 10- or 3-times greater than that produced by PEI-based previous method, respectively.We provide a rapid and efficient PEI-based method for preparation of recombinant lentiviral or retroviral vectors which is useful for making iPS cells as well as transduction of primary cell cultures.