【作者】 姚智慧； 祝晓春； 贺争鸣； 董关木； 卫礼； 巩薇； 吴惠英；

【Author】 YAOZhihui, ZHU Xiaochun, HE Zhengming, DONG Guanmu, Wei Li, GONG Wei, WU Huiying (The National Institute for the Control of Pharmaceutic & Biological Product Beijing 100050)

【机构】 中国药品生物制品检定所；

【摘要】 用sendai病毒E17株接种9日龄鸡胚尿囊腔,72h后收集尿囊液,用于提取病毒RNA,并逆转录成cDNA,用两对针对sendai病毒NP基因设计的外引物和内引物分别进行扩增.扩增产物克隆于T-载体,并测序.测序结果与Genbank报告的序列完全一致.尿囊液按10倍倍比稀释,进行敏感性实验.结果表明,第一次RCR可检测到10^-4病毒滴度,巢式PCR可检测到10^-7病毒滴度.将该方法用于检测用于生产疫苗的普通级乳地鼠肾和活疫苗OPV疫苗中的.sendai病毒,结果为阴性.更多还原

【Abstract】 The vims RNA was extracted from allantoic fluids of 9 days old chicken embryo inoculated with sendai virus, E17 strain, for 72 hours, and was reverse-transcribed into cDNA.Then cDNA was amplified by two primer sets respectively, designed according to the NP gene sequence of sendai virus.The PCR products were cloned into T - vector and se-quenced.The sequencing result was completely homology with the nucleotide sequence reported in Genbank.The sensitivity experiment was performed by serially diluting allantoic fluids, extracting RNA and then amplifying by RT- PCR.The result indicated that 10 ^-4 virus titer was detected by the first PCR with the outer primer sets and 10^-7 virus titer nested PCR with the inner primer sets. This method was used to detect sendai virus in the kidney of nurturing hamster, which was used for producing vaccine in China, and live vaccine, OPV.The results were negative.更多还原