西部马脑炎病毒基因组序列的RT-PCR检测  

Detection of the genomic sequence of western equine encephalitis virus by reverse transcription-PCR

作　　者：赵月峨[1] 于曼[1] 秦鄂德[1] 杨保安[1] 吕富双[1] 祝庆余[1] 

机构地区：[1]军事医学科学院微生物流行病研究所,北京100071

出　　处：《军事医学科学院院刊》2002年第3期200-202,共3页Bulletin of the Academy of Military Medical Sciences

摘　　要：目的 :建立敏感、特异的西部马脑炎 (westernequineencephalitis,WEE)病毒RT_PCR检测方法。方法 :首先采用逆转录法将病毒基因组RNA逆转录为cDNA ,然后以此cDNA为模板 ,进行PCR扩增。并对PCR扩增过程中的模板量、循环数、Mg2 + 浓度、退火温度和延伸时间等条件进行优化 ,以提高检测的特异性和敏感性。结果 :采用经优化的条件进行PCR扩增获得了约 35 0bp的单一DNA片段 ,其大小与预期的相一致 ,且可自 0 .1TCID50 的病毒液中检测出WEE病毒基因组序列。结论 :所建立的RT_PCR方法可自病毒感染的小鼠脑组织和传代细胞中检测WEE病毒的基因组RNA。Objective:To establish a specific and sensitive reverse transcription_PCR (RT_PCR) method for detection of the western equine encephalitis (WEE) virus genome.Methods:First, the viral genome RNA was transcripted to cDNA by reverse transcription. Then the fragment was amplified by PCR using the cDNA as template. In order to increase the specificity and sensitivity of RT_PCR, the conditions in PCR procedures, including the amount of template, the numbe of cycles, Mg 2+ concentration, annealing temperature etc, were optimized respectively.Results:A single and specific DNA fragment about 350 bp was amplified with the optimal PCR conditions. The size of the amplified fragment was equal to that of the expected product. The viral genome sequence in 0.1 TCID 50 of viruses can be detected.Conclusions:A specific RT_PCR assay was established for detecting the genomic RNA of WEE virus from infected suckling mice and culture cells.

关 键 词：西部马脑炎病毒 RT-PCR 基因组RNA 

分 类 号：R346[医药卫生—基础医学]