【摘要】 根据伪狂犬病病毒 (PRV)gB基因的序列 ,设计并合成了一对引物 ,以闽A株细胞培养毒为模板 ,筛选最佳反应条件 ,建立了检测PRV的PCR方法 应用该方法对Fb、Bartha、BJ、GD、V2F4、S、S3、SR、Buk、Shope、Norden、MinkⅢ、HB、F8、F9、F12等毒株的细胞培养液进行基因扩增 ,均获得了分子量为 2 81bp的特异性目的DNA片段 ,而对Vero细胞与FMDV、SVDV、HCV、PRRSV、JEV、PPV等病毒进行检测 ,结果均为阴性 ,没有出现交叉反应 对PRV毒株扩增的产物测序 ,结果序列与文献报道一致 ,证明PCR扩增产物和方法的特异性 对 1994～ 2 0 0 0年期间送检的临床样品和保存的PRV毒种 ,用病毒分离、双抗体夹心ELISA和PCR等 3种方法进行检测 ,结果前 2种方法检测为阳性的 ,PCR检测均为阳性 ;PCR检测为阴性 ,前 2种方法检测也为阴性 ;可是 ,前 2种方法检测为阴性的 ,PCR却检测出部分阳性 ;经x2 检验 ,证明PCR检出率明显高于前 2种方法的检出率 对PRV闽A株细胞毒提取物DNA进行检测 ,其最低检出量为 15 8pg 对 1999～ 2 0 0 0年期间广东、福建、海南等省的 31个大中型猪场送检的 191份病料进行检测 .结果病料阳性率为 2 6 2 % ( 50 191) ,猪场阳性率为 71% ( 2 2 31) 实验结果表明 ,所建立的PCR技术可用于伪狂犬病的快速诊断?更多还原

【Abstract】 One pair of primers that amplified the gB gene of pseudorabies virus (PRV)was designed and synthesized.PCR technique detecting the DNA of PRV was established after selecting the best reaction conditions.This technique was applied to specifically amplify the 281 bp DNA fragment of the PRV strains including Fa,Fb,Bartha,BJ,GD,V2F4,S,S3,SR,Buk,Shope,Norden,Mink Ⅲ,HB,F8,F9 and F12 in cultured samples.The negative results were achieved from Vero cells,swine vesicular disease virus,hog cholera virus,Japanese encephalitis virus,porcine reproductive and respiratory syndrome virus,porcine parvovirus,foot and mouth disease virus F29 strain,O3I3 strain,T509 strain and O Ⅱ MF249 strain.The results of sequencing showed that the PCR method was of specificity.The sensitivity of PCR reached 15.8 pg of PRV Fa strain DNA.The tissue samples obtained during 1994 and 2000 were detected,and the results showed that the sensitivity of PCR was more sensitive than virus isolation and the Sandwich ELISA.The PCR was applied to detect 191 tissue samples from 31 pig farms obtained from Guangdong,Fujian,Hainan Provinces during 1999 and 2000,50 samples(26.2%)were positive and 22 pig farms(71%)were positive.The results showed that established PRV PCR technique provided a more sensitive,specific and reliable method for diagnosis and epizootic study of the pseudorabies.更多还原