多重逆转录聚合酶链反应同步检测A组轮状病毒G基因型的研究  

One-step Simultaneous Detection of G-Genotype of Human Group A Rotaviruses by Multiplex RT-PCR

作　　者：唐少文[1] 叶临湘[1] 李燕[2] 王斌[2] 周玲[3] 康世秀[3] 杨继红[2] 

机构地区：[1]华中科技大学同济医学院流行病学教研室,武汉430030 [2]武汉市疾病预防控制中心 [3]武汉市儿童医院

出　　处：《中华检验医学杂志》2004年第4期234-236,共3页Chinese Journal of Laboratory Medicine

摘　　要：目的　为了检测A组轮状病毒VP7的基因型 ,建立一步多重逆转录聚合酶链反应 (RT PCR)同步检测A组轮状病毒G基因型技术。方法　在参照文献并分析A组轮状病毒VP7基因的基础上 ,选取了一个共用的下游引物RVG9和 6个A组轮状病毒G基因型G1、G2、G3、G4、G8和G9的特异性上游引物 ,利用多重RT PCR技术对标准株和样品进行检测。结果　进行一轮RT PCR ,即可至少同时检出 5种不同标准株混合物中的各个基因型。对纯化的A组轮状病毒标准株RNAO2 6 5进行系列稀释后 ,其检测灵敏度可达 1PFU。经对A组轮状病毒阳性的 6 0份样本进行检测 ,其中G3有 5 1例、G2有 4例 ,G1与G3混合感染 3例 ,另外尚有 2例不能分型。　结论　本研究建立的一步多重RT PCR技术对A组轮状病毒基因型研究将为临床诊断和实验研究提供一种快速、灵敏和特异的检测手段。Objective Simultaneously determination of G-genotype of human group A rotavirus. Methods 6 oligonucleotide primers were used,which were type-specific respectively for G1、G2、G3、G4、G8 and G9 G-genotype of human group A rotavirus.A multiplex polymerase chain reaction based assay was developed. The amplified product of 749bp, 652bp, 374bp, 583bp and 885bp represented genotype G1、G2、G3、G4 and G8 respectively. The concentrations of primers for multiplex PCR were optimized. Results All of five G-genotypes of human group A rotavirus could be typed simultaneously from a mixture of 5 standard strains.The sensitivity of the assay was 1PFU. 60 clinical samples showed that G3 was 85%, G2 was 6.7%. Conclusion The multiplex PCR appears to be a rapid, sensitive and specific method for detecting human group A Rotavirus′ genotype G and may be applied to clinical diagnosis and laboratory works.

关 键 词：多重逆转录聚合酶链反应 检测 轮状病毒G 基因型 

分 类 号：R446.5[医药卫生—诊断学]