携带GFP基因逆转录病毒感染原代培养脐带基质间充质细胞

摘要

目的 应用携带绿色荧光蛋白(GFP)基因的逆转录病毒感染原代培养脐带基质间充质细胞,为细胞重编程实验提供有效基因转移载体.方法 利用酶消化法在体外分离培养脐带基质间充质细胞.应用磷酸钙法将GFP基因转染到293T细胞内,包装生产携带GFP基因的逆转录病毒.收集病毒上清液感染脐带基质间充质细胞,应用倒置荧光显微镜观察逆转录病毒感染脐带基质间充质细胞情况.结果 体外分离培养出脐带基质间充质细胞; 293T细胞包装生产携带GFP基因逆转录病毒;包装生产后的逆转录病毒成功地感染脐带基质间充质细胞.结论 逆转录病毒可作为有效载体将外源性基因转染入脐带基质间充质细胞内,并且细胞能够稳定表达被导入的外源性基因.%Objective To explore efficiency of infecting umbilical cord matrix cells by retrovirus carrying GFP gene, to provide technical assurance for umbilical cord matrix cells reprogramming. Methods Umbilical cord matrix cells were isolated and cultured in vitro by enzyme digestion. GFP gene was transfected into 293T cells by applicating calcium phosphate transfection method, packaging and producing retrovirus with green fluorescent protein GFP gene. Umbilical cord matrix cells were infected by collecting virus supernatant, retroviral infection efficiency of umbilical cord matrix cells were observed through inverted fluorescence microscope. Results Obtained umbilical cord matrix cella in vitro. 293T cells successfully packaged and produced retrovirus carrying GFP. It was successful for the infection efficiency of umbilical cord matrix cell by packaging and producing retrovirus. Conclusions Exogenous gene can be transferred into umbilical cord matrix cells by retrovirus-mediated system, and the expression of exogenous gene is stable for the imported cells.