新型鸭呼肠孤病毒RT-PCR检测方法建立与初步应用

Establishment and Application of RT-PCR Method for Detection of NDRV

为建立一种能够快速检测新型鸭呼肠孤病毒(NDRV)的方法,本研究参考GenBank上登录的NDRV S3基因保守序列设计特异性引物。经条件优化后,建立了检测NDRV的RT-PCR方法。对其特异性、敏感性和重复性进行检验。结果显示:该方法仅能从NDRV分离毒中扩增到与预期大小相符,长度为298 bp的特异性目的片段,检测灵敏度达到83.4 pg病毒RNA;而其它病毒:番鸭呼肠孤病毒、禽呼肠孤病毒、鸭病毒性肝炎病毒、鸭瘟病毒、鸭新城疫病毒和禽流感病毒等样品的扩增结果均为阴性。采用该方法对在广东不同地区采集的15份鸭病料样品进行检测,NDRV阳性率为53.33%。表明建立的RT-PCR方法特异性强、敏感度高,可用于NDRV的临床诊断和流行病学调查。

To develop a method for detection of new-type duck reovirus （NDRV） , a RT-PCR method was established with 1 pairs of specific primers designed based on the conserved sequences of S3 gene ofNDRV. RT-PCR results showed that a 298 bp specifical fragment could be detected only from the RNA of NDRV-QY strain, and the sensitivity of RT-PCR reached to 83.4 pg NDRV RNA. Fifteen tissue samples of sick ducks from different areas of Guangdong province were detected respectively by the RT-PCR and the positive rate was 53.33% （8/15） for NDRV which indicated that the RT-PCR method for detecting NDRV was rapid, specific and sensitive, and could be used in clinic diganoses.