1型鸭肝炎病毒RT-PCR快速检测方法的建立

Establishment of RT-PCR Method for Detection of Duck Hepatitis Virus Type 1

为建立快速检测临床1型鸭肝炎病毒(DHAV-1)感染的方法,根据DHAV-1的vp1基因序列设计1对引物,以DHAV-1及其他常见的感染鸭的病毒基因组为模板,建立了DHAV-1的特异性RT-PCR检测方法;以10倍梯度稀释的DHAV-1 RNA为模板,测定该方法的灵敏度;采集江苏多地的疑似DHAV-1感染病料,提取基因组进行RT-PCR扩增,产物经测序鉴定后判断所建立方法的检测率。结果显示,建立的方法可以特异性扩增DHAV-1 vp1基因保守区360 bp的序列,最低可检测1 fg基因组模板,对临床样品检测率为100%。表明,成功建立了特异性强、灵敏度高且可以用于临床快速诊断DHAV-1感染的方法。

In order to establish a rapid detection method for dicnical duck hepatitis virus type 1(DHAV-1)infection,a pair of primers based on vp1 gene sequence was designed.Using DHAV-1 or other common disease virus genome as template,a specificity RT-PCR method was established.The template was ten times diluted for sensitivity detection.Some suspected DHAV-1 infected epidemic materials were collected from Jiangsu province.Genome of the materials were extracted and detected followed with sequence determination.The results showed that the established method could specifically amplify vp1 conservative region 360 bp sequence.The sensitivity and specificity results showed that the detection limitation was1 fg,and there was no cross reactions with other duch virused.The detection rate of clinical samples was100%.The results showed that a high specificity and high sensitivity RT-PCR method for rapid clinical diagnosis of duck DHAV-1 infection was successfully established.