鸡传染性支气管炎病毒M41型的扩繁及在鸡胚肾细胞上的适应性研究

摘要

探明鸡传染性支气管炎病毒(Avian Infectious Bronchitis Virus,IBV)感染鸡胚肾细胞(Chicken Embryo Kidney Cells,CEK cell),并且实现其增殖为目的.将IBV稀释液注射到SPF鸡胚尿囊腔内,培养鸡胚扩繁,观察记录结果;并培养CEK细胞,采用100倍半数组织培养感染量的IBV进行染毒实验;应用Reed-Muench法计算半数鸡胚感染量为10-6.68/0.1 mL;用Real time PCR检测IBV在感染细胞中的含量,以确定IBV在CEKcells中已经感染且增殖成功.结果表明,接种10日龄的SPF鸡胚可以收集更多的、约8 mL的尿囊液毒;反复盲传至7代的IBV可使CEK cells产生明显的细胞病变效应;通过实时荧光定量PCR和RT-PCR检测,最终确定IBV-M41型扩繁成功并且能够在CEK细胞上繁殖,为下一步实验提供了参考.%This study investigated the avian infectious bronchitis virus (IBV) - M41 infected chicken embryo kidney cells (CEK cell) and could multiply successfully . The IBV was injected into the SPF chicken eggs to propagate. And CEK cells were cultured ahead of adding in the IBV of 100TCID50. Reed-Muench was used to calculate the EIDso . The real time PCR assay was used to detect the content of IBV in CEK cells whitch past infection. IBV-M41 was determined and it multiplied in CEK cells successfully. The results showed that the 10-day-old SPF chicken embryos inoculated can collect more of allantoic fluid virus(a-bout 8 mL) ; the CEK cells infected with 7th generation of IBV can produce significant cytopathic effect; and ultimately determined by quantitative real-time PCR and RT-PCR assay, the propagation of IBV-M41 and the ability to reproduce in CEK cells was succesfful. The results of this study provides a reference for further experiments.