| 摘要: |
| 为了优化猪瘟病毒的RT-nested PCR检测方法,对福建省猪瘟流行情况进行了调查。根据GenBank上发表的猪瘟病毒Shimen株基因序列设计并合成了2对引物,优化了猪瘟病毒(Classical swine fever virus,CSFV)的RT-nested PCR检测方法,并对所优化的RT-nested PCR特异性进行了检验。结果表明,该方法检测CSFV cDNA含量的最低极限为1×10-7ng/mL,只有CSFV扩增出了272 bp的目的条带,从猪繁殖与呼吸综合征病毒(PRRSV)、猪流感病毒(SIV)、猪圆环病毒2型(PCV2)、猪伪狂犬病病毒(PRV)和猪细小病毒(PPV)阳性毒和健康猪脾、肝组织及阴性PK-15细胞均未扩增出特异性条带,说明该方法特异性强。应用此方法对福建省133份病料进行检测,结果有60份病料为阳性,阳性率为45.1%。结果提示优化的检测方法灵敏度高,特异性强;福建省猪群CSFV感染率高,需要加强CSF预防控制工作。 |
| 关键词: 猪瘟病毒 RT-nested PCR 检测方法 |
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| Optimization and application of RT-nested PCR detection for classical swine fever virus |
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| Abstract: |
| In order to optimize of RT-nexted PCR detection method for classical swine fever virus,the prevalence of classical swine fever in FuJian province was investigated.Two pairs of primers were designed and synthesized to test the sensitivity and specialization by referring to strain Shimen,and the method of detection for CSFV was optimized.The result of sensitivity showed that the most dilution cDNA of CSFV was 1×10-7 ng/mL,and the method could not amplify any band from PRRSV,SIV,PCV2,PRV,PPV spleen and liver of health pig and negative control PK-15 cell except CSFV,which indicated that the RT-nested PCR method was special for CSFV.When this method was applied to detect 133 samples swine tissue from Fujian Province,60 samples were positive,and the positive ratio was 45.1％.The result showed that the RT-nested PCR method was of high sensitivity and specialness;the infection ratio of swinery was high in Fujian Province,and it was needed to strengthen prevention and control of CSF work. |
| Key words: classical swine fever virus RT-nested PCR detection method |
