中国组织工程研究2017，Vol.21Issue(29)：4642-4647,6.DOI:10.3969/j.issn.2095-4344.2017.29.009
三种携带GFP慢病毒转染兔骨髓间充质干细胞的效果
Transfection efficiency of three different lentiviruses with green fluorescent protein gene to transfect rabbit bone marrow mesenchymal stem cells
摘要
Abstract
BACKGROUND:We explore the transfection effects of different lentiviruses on bone marrow mesenchymal stem cells (BMSCs) from the same source in order to screen out appropriate cell transfection tools.OBJECTIVE:To explore the difference in the transfection efficiency of three different lentiviruses carrying green fluorescent protein (GFP) to transfect the rabbit BMSCs.METHODS:Bone marrow samples were extracted from the ilium of 2-week-old Japan white rabbits,and then the BMSCs were separated,purified and extended using density gradient centrifugation.Cell surface markers were identified using flow cytometry.pTomo-GFP lentivirus,Ad-GFP lentivirus and lenti-GFP lentivirus were used to transfect the BMSCs,and the transfection efficiency was tested using flow cytometry.The cell viability was detected using cell counting kit-8 before and after transfection.RESULTS AND CONCLUSION:As the positive rates of CD44,CD34 and CD45 were 75.64％,6.14％ and 9.48％,respectively,the cultured cells were identified as BMSCs by the flow cytometry.The expression levels of GFP after transfection by pTomo-GFP,Ad-GFP and ienti-GFP were 2.64％,16.72％ and 45.24％,respectively.The cell growth curve showed that pTomo-GFP lentivirus,Ad-GFP and ienti-GFP lentivirus showed certain effects on cell growth,pTomo-GFP and Ad-GFP had an effect on cell growth at incubation period and logarithmic growth phase at 1-8 days after transfection,which may cause some interference to the viability of early cells,ienti-GFP had little effect on cell growth in the short term,but there was a change in cell growth at day 12 after lenti-GFP transfection,and a long-term effect of lenti-GFP on cell viability was considered.Therefore,lenti-GFP lentivirus could highly transfect the BMSCs and could not impact the normal growth of BMSCs,indicating that lenti-GFP lentivirus is suitable for BMSCs tracing and gene transfection.关键词
干细胞/骨髓干细胞/绿色荧光蛋白/间充质干细胞/骨髓间充质干细胞/慢病毒/转染/转染率/生长曲线/国家自然科学基金分类
医药卫生引用本文复制引用
肖渝,李彦林,王坤,李龙滕,闫祥甲,蒙旭晗..三种携带GFP慢病毒转染兔骨髓间充质干细胞的效果[J].中国组织工程研究,2017,21(29):4642-4647,6.基金项目
国家自然科学基金(81460340) （81460340）
云南省骨关节疾病诊疗省创新团队项目(2014HC1018) （2014HC1018）
医疗卫生内设机构研究项目(2014NS161) （2014NS161）
the National Natural Science Foundation of China,No.81460340 （）
the Yunnan Provincial Innovation Team Project for Diagnosis and Treatment of Bone and Joint Disease,No.2014HC1018 （）
the Research Project of Medical and Health Institutions,No.2014NS161 （）
