【摘要】 建立了一种快速检测HTLV-I前病毒的荧光定量方法。利用HTLV前病毒DNA序列设计特异性TaqmanMGB探针及引物对目标DNA片段进行实时检测,用含目标片段质粒构建标准曲线。该定量检测方法在103~107copy/μL范围内与Ct值呈良好线性关系(R2可达0.999),最低检测浓度为5copy/μL。利用该系统对献血者血液标本进行HTLV前病毒初筛确认及定量检测,结果表明该方法重复性良好,特异性高,适用于献血者和临床标本的检测和确认,亦可用于对HTLV病毒载量与成人T细胞白血病等疾病的关联性进行研究。更多还原

【Abstract】 A quantitative real-time PCR assay was developed to measure the proviral load of human T-lymphotropic virus type I(HTLV-I) in peripheral blood.The technology utilizes special primers and Taqman MGB fluorescence probe to measure amplification products from the gag-pro-pol polyprotein gene of HTLV-I.HTLV-I copy number was normalized to the amount of cellular DNA by quantitation of the β-actin gene.The amplification system was sensitive to detect 5copy/μL.The standard curve had a good linearity when the quantity for the gene was between 103 and 107 copy/μL(R2=0.999).Good reproducibility was observed in each intra-and inter-assay.We also measured proviral load in peripheral blood in 12 HTLV-I seropositive former blood donors.Proviral load for HTLV-I infected donors ranged from 0.015 to 12.819 copy/cell in WBC with the mean of 3.116 copy/cell.更多还原