多聚赖氨酸氧化铁纳米粒制备与体外转染实验研究

Preparation of poly-L-lysine iron oxide nanoparticles and in vitro experiment

目的评价多聚赖氨酸氧化铁磁性纳米粒作为基因载体在体外转染中的可行性。方法应用改进的共沉淀法制备多聚赖氨酸氧化铁磁性纳米粒(poly-L-lysine iron oxide nanoparticles,PLL-IONP)。使用透射电镜检测PLL-IONP的形态和粒径,采用琼脂糖凝胶电泳分析PLL-IONP结合质粒pEGFP-C1及抵抗DNaseⅠ消化的能力。以PLL-IONP作为基因载体将质粒pEGFP-C1转染入SMMC-7721细胞中。结果 PLL-IONP分散性好,大小均一,粒径在35nm左右。琼脂糖凝胶电泳显示,PLL-IONP在各种质量比均有良好的DNA结合能力。DNA保护实验显示PLL-IONP能有效保护DNA不被DNaseⅠ降解。荧光显微镜观察到了绿色荧光蛋白在SMMC-7721细胞中表达。结论成功制备了多聚赖氨酸氧化铁磁性纳米粒,该纳米粒在体外可作为基因载体,有效的介导质粒pEGFP-C1转染。

【Objective】To evaluate feasibility of the poly-L-lysine iron oxide nanoparticles（PLL-IONPs） as gene carrier in vitro.【Methods】The PLL-IONPs were synthesized with improved co-precipitation method.The configuration and diameter of PLL-IONPs were detected using transmission electron microscope.The potential of adsorbing plasmid pEGFP-C1 and resisting DNaseⅠ digestion of PLL-IONPs was analyzed by agarose gel electrophoresis.PLL-IONPs bound with plasmid pEGFP-C1 as one of gene carriers were transfected into human hepatocellular carcinoma cell line SMMC7721.【Results】These compound nanoparticles were very uniform and dispersed homogeneously.Under zetasizer,the mean diameter of the PLL-IONPs was 35 nm.Agarose gel electrophoresis of binding experiment and protection experiment showed that PLL-IONPs had satisfactory potential to adsorb DNA and to protect DNA from DNaseⅠ digestion.The expression of green fluorescent protein was observed by inverted microscope.【Conclusions】PLL-IONPs have been prepared successfully.It can transfect the plasmid into human hepatocellular carcinoma cell line SMMC7721 in vitro.