阳离子脂质体介导肽核酸转染K562细胞的研究

Experimental study on peptide nucleic acid transfection into K562 cells mediated by cationic liposome

目的优化阳离子脂质体介导肽核酸(PNA)转染K562细胞的条件。方法以阳离子脂质体lipofectamine 2000为载体,将标记有FITC荧光基团的PNA转染K562细胞,在荧光显微镜下观察并计算其转染效率,采用Cell Counting Kit(CCK-8)检测其细胞毒性。应用正交试验筛选出最佳的PNA和脂质体的用量及比例,并优化稀释用培养基血清浓度,以获得最优的转染效果。结果以2.5×105/mL^3×105/mL的细胞密度接种,100μL/孔的培养基中加入PNA 6.25pmol,PNA与脂质体的体积比为1∶3.5,稀释用培养基未加胎牛血清时,细胞转染效率和细胞毒性最佳,转染效率为87.2%,细胞存活率(RGR)为94.1%。结论PNA可通过阳离子脂质体转染进入K562细胞,优化条件后得到的细胞转染效率和细胞存活率能够满足基因表达研究的实验要求。

Objective Abstract Objective To optimize the transfection conditions for peptide nucleic acid（PNA）into K562 cells mediated by cationic liposome.Methods With the cationic liposome lipofectamine 2000 used as vector,PNAs marked by FITC fluorophore were transfected into K562 cells.Transfection efficiency was calculated according to the observation of cells under fluorescence microscope,and cytotoxicity was examined using Cell Counting Kit（CCK-8）.In order to achieve the best transfection results,the dose and proportion of PNA and liposomes were optimized by orthogonal experiment,as well as the serum concentration in medium used as dilution was optimized.Results When the cells with the density of 2.5×105/mL-3×105/mL were cultured,the optimized transfection conditions were as follow：each 100μl medium added with 6.25 pmol PNA,the volume proportion of PNA to liposomes was 1∶3.5,and the medium used as dilution had no fetal bovine serum.Under these conditions,the best transfection results were achieved,with a transfection efficiency of 87.2%and a relative growth rate（RGR）of 94.1%.Conclusion PNA could be transfected into K562 cells mediated by cationic liposome.The cells transfection efficiency and RGR achieved under the optimized transfection conditions could satisfy the requirement of gene expression research.