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目的:观察腺相关病毒载体介导外源基因导入人外周血干细胞的转染效率及表达。方法:本文采用腺相关病毒(adeno-associated virus,AAV)载体介导的基因转移方法,将绿色荧光蛋白(green fluorescent protein,GFP)基因和人多药耐药基因(multi-drug resitance,MDR1)导入人外周造血干/祖细胞,并对转染效率和转基因造血细胞的耐药性进行了初步研究。结果:rAAV/GFP感染CD34阳性细胞后,在荧光显微镜下观察,约30％细胞中可见绿色荧光。将rAAV/MDR1重组病毒感染CD34阳性细胞,经PCR和MTT法证实,导入MDR1基因的CD34阳性细胞对秋水仙素的耐药性与未导入MDR1基因的细胞有明显差异。结论:腺相关病毒载体能有效地将外源基因导入外周血CD34阳性细胞,并可在其中有效地表达。
OBJECTIVE: To observe the transfection efficiency and expression of foreign gene into human peripheral blood stem cells mediated by adeno-associated virus vector. Methods: A green fluorescent protein (GFP) gene and a multi-drug resitance (MDR1) gene were transfected by adenovirus vector-mediated gene transfer (AAV) The human hematopoietic stem / progenitor cells were introduced into human and the transfection efficiency and the drug resistance of transgenic hematopoietic cells were studied. Results: After being infected with rAAV / GFP, CD34 positive cells were observed under fluorescence microscope and green fluorescence was observed in about 30% of the cells. The CD34 positive cells were infected with rAAV / MDR1 recombinant virus. The results of PCR and MTT confirmed that the CD34 positive cells transfected with MDR1 gene had significant differences in the resistance to colchicine compared with the cells without MDR1 gene. CONCLUSION: Adeno-associated virus vectors can efficiently transduce exogenous genes into peripheral blood CD34-positive cells and can be efficiently expressed in them.