磁捕法对传染性非典型肺炎冠状病毒RNA富集体系的初步研究  

The detection of SARS-associated coronavirus by a specific RNA-capture polymerase chain reaction

作　　者：黑爱莲[1] 蔡剑平[2] 汪仕奎[2] 胡继红[2] 张传宝[2] 申子瑜[2] 郭健[2] 

机构地区：[1]中国医学科学院中国协和医科大学,北京100730 [2]中国医学科学院卫生部临床检验中心,北京100730

出　　处：《中华检验医学杂志》2004年第5期286-288,共3页Chinese Journal of Laboratory Medicine

基　　金：国家高技术研究发展计划(863计划)基金资助项目(2002AA215013);中央保健委员会办公室资助项目(0302)

摘　　要：目的 初步研究建立磁捕法对传染性非典型肺炎又称严重急性呼吸综合征(SARS)冠状病毒RNA的富集、纯化体系,以提高逆转录聚合酶链反应(RT-PCR)检测SARS冠状病毒RNA的敏感性。方法 根据国际基因库公布的SARS冠状病毒基因序列,自行设计引物、探针,体外克隆目的RNA片段作为标准物质;利用标准RNA以及模拟SARS血清标本的RNA,评价自行设计的SARS冠状病毒巢式RT-PCR检测体系和磁捕法SARS冠状病毒RNA富集检测体系的检测效果。结果SARS冠状病毒巢式RT-PCR检测体系,对于模拟血清SARS标本RNA的最低检测样本浓度为20拷贝/μl,而磁捕法SARS冠状病毒RNA富集检测体系的最低检测样本浓度为1拷贝/μl。结论 初步建立的磁捕法SARS冠状病毒RNA富集检测体系,可有效地对低浓度的RNA样本进行浓缩、纯化,有效提高SARS RT-PCR检测方法的敏感性。Objective The enrichment for severe acute respiratory syndrome ( SARS)-associated eoronavirus RNA by capturing RNA onto magnetic beads was investigated to improve the sensitivity of reverse transcriptase-polymerase chain reaction (RT-PCR) method. Methods Primers and probes were designed according to the published sequence of the SARS-associated eoronavirus gene. The specific RNA fragment was transcribed in vitro and was quantitied as a standard to evaluate the sensitivity of the detection asssays. Results The minimum concentration of the RNA sample detected by the routine nested RT-PCR assay was 20 copies/μl, however, the minimum concentration of the RNA sample detected by the RNA-capture nested RT-PCR assay was 1 copies/μl. Conclusion The SARS-associated eoronavirus nested RNA-capture RT-PCR method was initially established with higher sensitivity than routine nested RT-PCR method, due to the enrichment for the SARS-associated eoronavirus RNA.

关 键 词：磁捕法 传染性非典型肺炎 冠状病毒 RNA富集体系 

分 类 号：R511.9[医药卫生—内科学]