鉴别猪伪狂犬病病毒的双重PCR方法的建立及初步应用

doi:10.16431/j.cnki.1671-7236.2016.05.007

Abstract

Abstract: In order to detect the procine pseudorabies virus (PRV) infection and differentiate virulent and avirulent PRV, a duplex PCR method was established with two pairs of specific primers designed based on the conserved sequences of gE and gB genes of PRV.Optimize the reaction conditions by adjusting concentration of primers from 0.2 to 1.4 μL, each time adding 0.2 μL and temperature of annealing from 56 to 60 ℃, added 1 ℃ each time, and so on.The results showed that the suitable annealing temperature was 56 ℃ and the suitable primer dose was 1 μL.Two special fragments of 316 bp (PRV gE) and 432 bp (PRV gB) were amplified by the optimized duplex PCR.The specific tests showed that no PCR products were detected for PCV2, PTV, CSFV, PPV, PRRSV and E.coli.The sensitivity test showed that DNA used for the duplex PCR might be as low as 4.4×10³ copy/μL for PRV gE and 3.3×10³ copy/μL for PRV gB.The sensitivity of the duplex PCR was close to the single PCR.Fifty-six clinical samples of sick pigs from different areas in Guangdong and Guangxi province were detected by the duplex PCR and the virulent detection rate was 53.6% (30/56) for PRV and the negative rate was 46.4%(26/56), and there was no attenuated PRV infection.

Key words: porcine; pseudorabies virus; duplex PCR; detection method

CLC Number:

S852.65

LIAN Si-nan, LIANG Lin, BAI Ai-quan, CUI Shang-jin. Establishment and Primary Application of Duplex PCR Assay to Detect Procine Pseudorabies Virus[J]. , 2016, 43(5): 1169-1175.

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Establishment and Primary Application of Duplex PCR Assay to Detect Procine Pseudorabies Virus

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