实时荧光定量PCR技术在SPF鸡四种垂直传播疾病监测中的应用

目的探讨荧光定量PCR检测技术对SPF鸡四种垂直传播病毒的检测应用。方法采集60份SPF鸡及70份普通鸡群蛋清、泄殖腔试子样品,提取样品核酸,分别进行ARV、REV、CAV、ALV四种病毒实时荧光定量PCR检测,根据标准曲线及溶解曲线分析判读样品病毒拷贝数。结果 SPF鸡ALV 2份阳性,检出率3.3%,其余病毒检测均为阴性;普通鸡样品REV检测2份阳性,检出率2.9%,ALV 10份阳性,检出率14.3%。结论荧光定量PCR检测方法最低可检测到100个拷贝核酸,检测灵敏度较高,有望应用于SPF鸡临床样品的病原检测。

Application of SYBR Green Fluorescent Quantitative Real-Time PCR in the Detection of Four Vertical Transmission Viruses in SPF Chickens

Objective To apply SYBR green I fluorescent quantitative real-time PCR（QPCR） in detecting 4 vertical transmission viruses of chicken and to evaluate its application value for the quality control of SPF chickens.Method Quantitative real-time PCR technique was used to detect REV,ALV,ARV and CAV RNA,DNA or provirus DNA in 30 cloacal swab samples and 30 eggs of SPF chicken samples as well as 40 cloacal swab samples and 30 eggs of common chicken samples.Standard curve and melting curve analyses were conducted to obtain the quantitative result.Results Out of the SPF chicken samples,2/30（3.3%） were ALV-positive,and the three other viruses were negative.For the common chicken samples,2/70（2.9%） were REV-positive,and 10/70（14.3%） were ALV-positive.Conclusions These QPCR assays show better sensitivity and specificity for detecting the 4 viruses in SPF chickens with a detection limit of 100 copies of target DNA per-reaction,and have a bright prospect in clinical application.