Abstract
Objectives
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.
Results
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 °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.
Conclusion
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.
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Acknowledgments
This work was supported by the National Nature Science Foundation of China (No. 81571994, 81570567), the Natural Science Foundation of Guangdong province, China (No. 2014A030313483, 2015A030313447), and the Scientific Research Foundation for the Returned Overseas Chinese Scholars, State Education Ministry. We would like to thank Dr. Stanley Lin for kindly revising this manuscript.
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The authors Shaozhe Yang and Haijun Shi have contributed equally to this work.
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Yang, S., Shi, H., Chu, X. et al. A rapid and efficient polyethylenimine-based transfection method to prepare lentiviral or retroviral vectors: useful for making iPS cells and transduction of primary cells. Biotechnol Lett 38, 1631–1641 (2016). https://doi.org/10.1007/s10529-016-2123-2
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DOI: https://doi.org/10.1007/s10529-016-2123-2