一种高效稳定的磷酸钙转染293T细胞方法的建立及评价

doi:10.13481/j.1671-587x.20190535

Abstract

Abstract: Objective:To explore the effects of two kinds of calcium phosphate transfection methods in the 293T cells, and to establish the method of achieving high-efficiency and stable calcium phosphate transfection in the 293T cells. Methods:Fluorescence microscope was used to observe the transfection efficiencies of transfection of pCDH-GFP-3xflag-TRAF6 plasmid into the 293T cells by two kinds of calcium phosphate transfection methods (traditional calcium phosphate transfection method and improved calcium phosphate transfection method). Real-time PCR and Western blotting method were used to detect the expression levels of TRAF6 mRNA and flag protein in the 293T cells after transfection of pCDH-GFP-3xflag-TRAF6 plasmid by two kinds of calcium phosphate transfection methods. Results:Under fluorescence microscope, compared with traditional calcium phosphate transfection method, the transfection efficiencies of improved calcium phosphate transfection method 24 an 48 h after transfection of pCDH-GFP-3xflag-TRAF6 plasmid into the 293T cells were significantly increased (P<0.01). The Real-time PCR and Western blotting results showed that compared with traditional transfection method, the expression levels of TRAF6 mRNA and flag protein in the 293T cells 24 and 48 h after transfection of pCDH-GFP-3xflag-TRAF6 plasmid by calcicum phosphate transfection method were significantly increased(P<0.01). Conclusion:The improved calcium phosphate transfection method established in this reseach is a highly efficient and stable DNA transfection method.

Key words: calcium phosphate transfection, 293T cells, tumor necrosis factor receptor associated factor 6, flag protein

CLC Number:

Q819

HUA Jin, CHENG Zhibin, LIN Chunlin, DAI Qibao, ZHU Guangwei. Establishment and evaluation of a highly efficient and stable calcium phosphate transfection method for 293T cells[J].Journal of Jilin University(Medicine Edition), 2019, 45(05): 1177-1181.

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Establishment and evaluation of a highly efficient and stable calcium phosphate transfection method for 293T cells

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