為建立一種快速、準(zhǔn)確,、常規(guī)的豬血凝性腦脊髓炎病毒（porcine hemagglutinating encephalomyelitis virus,，PHEV）檢測方法，根據(jù)PHEV N蛋白基因序列,，設(shè)計(jì)１對引物和１條探針,，建立了PHEV實(shí)時(shí)熒光RT-PCR檢測方法。該方法特異性好,，與豬繁殖與呼吸綜合征病毒（PRRSV）,、豬偽狂犬病病毒（PRV）、豬流行性腹瀉病毒（PEDV）,、豬傳染性胃腸炎病毒（TGEV）,、豬瘟病毒（CSFV）和豬圓環(huán)病毒2型（PCV2）均無交叉反應(yīng)；用陽性質(zhì)粒Puc57-PHEVn評估其敏感性,，發(fā)現(xiàn)檢測下限達(dá)10-6 ng/μL（224拷貝/μL）,。采集233份發(fā)病豬組織樣品進(jìn)行臨床檢測，發(fā)現(xiàn)該方法與巢式RT-PCR的符合率達(dá)98.3%,，準(zhǔn)確篩查出巢式RT-PCR并測序陽性的樣品14份,，且檢出率高于巢式RT-PCR。結(jié)果表明,，本方法敏感,、特異，可用于PHEV臨床樣品的檢測,。

Establishment and Application of a Real-time RT-PCR Assay for Detection of

Porcine Hemagglutinating Encephalomyelitis Virus

In order to establish a rapid,，accurate and routine method for detection of porcine hemagglutinating encephalomyelitis virus（PHEV），a pair of primers and a probe were designed based on the N gene sequence of PHEV,，and a real-time RT-PCR assay was developed,，which was with good specificity，and failed to crossly react with porcine reproductive and respiratory syndrome virus（PRRSV）,，porcine pseudorabies virus（PRV）,，porcine epidemic diarrhea virus（PEDV），transmissible gastroenteritis virus（TGEV）,，classical swine fever virus（CSFV）and porcine circovirus type 2（PCV2）. The sensitivity of the method was evaluated based on positive plasmid Puc57-PHEVn,，and it was found that the limit detection was 10-6 ng/μL（224 copies/μL）. A total of 233 swine tissues were tested by the method,，and it was found that 14 positive samples were detected out，and the coincidence rate with the nested RT-PCR was 98.3%,，but the detection rate was higher than that by nested RT-PCR. It was concluded that the method could be used to detect PHEV clinical samples benefited from its high sensitivity and specificity..

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