摘要
目的:建立慢病毒-绿色荧光蛋白载体有效感染人包皮成纤维细胞的方法,为利用慢病毒载体介导外源基因导入人包皮成纤维细胞,后者直接诱导分化为多能干细胞的研究奠定基础。方法:采用0.1%Ⅰ型胶原酶冷消化联合组织块贴壁法培养人包皮成纤维细胞,在细胞培养液中分别加入不同感染复数的慢病毒和具有促进病毒感染的细胞增强液,并于感染48h后观察慢病毒转导情况。结果:成功建立人包皮成纤维细胞培养模型,当慢病毒对人包皮成纤维细胞的感染复数为20及在培养基中加入细胞增强液时,感染效率可以达到80%,能满足后续实验的需要。结论:慢病毒较易感染人包皮成纤维细胞,是具有发展潜力的多能干细胞细胞技术研究的新载体。
Objective To investigate the effective and stable transduction of human foreskin fibroblast by lentivirus vector,which can provide a basis for induced pluripotent stem cell research.Methods Human foreskin fibroblast was isolated and cultured with the technique of 0.1% collagenase I digesting.Lentivirus with green fluorescent protein infected cells with three gradients were added into DMEM.Three groups of cells with multiply of infection were added respectively enhanced infection solution after being infected for 48 hours.The expression of green fluorescent proteins and infection efficiency were observed with inverted microscope.Results The model of human foreskin fibroblast was successfully established.The best multiply of infection for lentivirus infecting human foreskin fibroblast was 20,which could achieve 80% infection efficiency.However,enhanced infection solution a kind of infected reagents of polycation was need added in DMEM in order to improve the infection efficiency of lentiviral vectors.Conclusion Lentiviru is easy to infect human foreskin fibroblast and demonstrates a high infection efficiency of human foreskin fibroblast,which is a new vector for iPS technology research.
出处
《中华实用诊断与治疗杂志》
2009年第12期1194-1196,共3页
Journal of Chinese Practical Diagnosis and Therapy
基金
福建省条件建设项目(2005Q006)
福建省科技厅社会发展重点项目(2009Y0009)
关键词
慢病毒载体
皮肤成纤维细胞
多能干细胞
Lentivirus vector human foreskin fibroblast induced pluripotent stem cell
