聚合酶链反应技术检测禽网状内皮组织增殖病病毒

目的建立聚合酶链反应(PCR)技术检测禽网状内皮组织增殖病病毒(REV)的方法。方法提取感染REV-T和脾坏死病毒(SNV)的SPF鸡胚成纤维细胞DNA为模板,利用前病毒长末端重复序列(LTR)区引物进行扩增。采集肿瘤病鸡,以及人工感染REV 28 d后鸡肝脏、脾脏、肾脏、心脏、胸腺、法氏囊等器官,进行扩增。同时将采集的脏器组织,进行HE染色和免疫组化试验(IHC)。结果REV-T感染的组织未检测出电泳条带,而SNV感染的细胞中检测到了一条300bp特异而清晰的电泳条带,而且SNV感染的鸡组织中,PCR方法检测到了特异的条带。通过HE染色和免疫组化技术观察到了肿瘤组织,肿瘤细胞的形态、分布。结论PCR检测REV更快捷,特异更好。

Detection of Reticuloendotheliosis Virus by Polymerase Chain Reaction Technique

Objective To establish a method of polymerase chain reaction(PCR) for detection of reticuloendotheliosis virus(REV).Methods The genomic DNA extracted from SPF chick embryo fibroblasts(CEFs) infected with REV-T and SNV was used as the template,followed amplification by primers from the long terminal repeats(LTR) of provirus.Consequently,the tissues from tumor-bearing chickens,as well as liver,spleen,kidney,heart,thymus and bursa from chickens on day 28 after having artificially been inoculated REV,were collected.Parts of the whole tissues were subjected to DNA extraction for PCR.The others were stained with haematoxylin and eosin (HE) and also immunohistochemical staining(IHC).Reslts Nothing amplified from REV-T was detected on agarose gel by electrophoresis.A specific and clear band sized about 300 bp was showed after amplification from SNV.Moreover,specific bands were detected by PCR of the tissues of birds infected with SNV.The tumor tissues,appearance and distribution of tumor cells were observed by histopathology(HE and histochemical staining).Conclusion The PCR is a specific and rapid technique for detection of reticuloendotheliosis virus.