逆转录病毒载体介导的双耐药基因转染小鼠骨髓细胞的实验研究

目的:将耐药基因以逆转录病毒为载体转染骨髓细胞以解决大剂量化疗所造成的骨髓抑制。方法:以逆转录病毒为载体,将双突变的二氢叶酸还原酶基因(DH-FR)和胞苷脱氨基酶基因(CD)转染小鼠骨髓细胞,观察该细胞耐MTX及Ara-C的CFU-GM生成情况;RT-PCR检测转基因的表达情况。结果:转基因的小鼠骨髓细胞有耐药克隆形成(14%);基因转染后小鼠骨髓细胞对MTX和Ara-C的耐受明显增加,P<0·005;RT-PCR显示转基因细胞有转染的基因条带。结论:双耐药基因通过逆转录病毒转染小鼠骨髓细胞并且获得表达,提高了骨髓细胞对MTX和Aar-C的耐药性。

Experimental study on mice bone marrow cells transferred with double-drug resistant gene by retroviral vectors

OBJECTIVE:To overcome the myelosupperssion caused by high dose chemotherapy by transfering drug resistance gene into bone marrow cells. METHODS: Double-mutant dihydrofolate reductase-cytidine deaminase gene was transferred into mouse bone marrow cells by retroviral vector. Colony-forming unit granulocyte-macrophage (CFU-GM) to MTX and Ara-C was gotten by cell culture. The expression of transferred drug resistance gene in mouse bone marrow cells was examined by RT-PCR. RESULTS: Yield of CFU-GM of transfected mouse bone marrow cells was 14% and its drug resistance to both MTX and Ara-C was increased obviously, P<0.005. The expression of drug resistance gene in transfected mouse bone marrow cells was demonstrated by RT-PCR. CONCLUSIONS: Double drug resistance gene can not only integrate and express in mice bone marrow cells by retroviral vector but also increase the drug resistance to MTX and Ara-C.