鉴别非洲猪瘟病毒和猪瘟病毒野毒株二重TaqMan MGB实时荧光定量PCR检测方法的建立及初步应用

doi:10.11843/j.issn.0366-6964.2021.018

Abstract

Abstract: To detect the African swine fever virus (ASFV) and Classical swine fever virus (CSFV) wild strains rapidly, a specific and sensitive real-time fluorescence quantitative PCR (FQ-PCR) method was established by using the TaqMan MGB probe technique. Specific primers and probes were designed based on the P72 gene of ASFV and 5' non-coding region of the CSFV wild strain to establish the duplex FQ-PCR assay based on TaqMan MGB probe technique. The sensitivity, specificity and stability of FQ-PCR were determined, and 50 clinical samples were simultaneously detected by the FQ-PCR assay, the national standard of CSFV and the OIE recommended detection method for ASFV. The results indicated that the duplex FQ-PCR assay was successfully established and showed a good linear relationship at a template range of 10⁰-10⁶ copies·μL^-1; the specificity of duplex FQ-PCR assay revealed that amplifications were showed synthetic on ASFV and CSFV genes, but other pathogens have no amplifications; variation coefficient of intra and inter assay were 1.18%-2.08%, respectively; the sensitivity of the assay was 10 copies·μL^-1; Fifty clinical samples was detected by the duplex FQ-PCR, the results was consistent with detection of CSFV by national standard and detection of ASFV by OIE recommended method. The duplex TaqMan MGB FQ-PCR assay of ASFV and CSFV wild strain was specific, sensitive, rapid and suitable for identify detection of ASFV and CSFV wild strains.

Key words: African swine fever virus, classical swine fever virus wild strains, duplex TaqMan MGB FQ-PCR, detection

CLC Number:

WANG Shujuan, BAN Fuguo, WANG Dongfang, LIU Ying, ZHAO Xueli, XIE Caihua, WANG Cui, MA Zhenyuan, YANG Haibo, CHAI Mao, YAN Ruoqian. Establishment and Application of a Duplex TaqMan MGB FQ-PCR for Differential Detection of African Swine Fever Virus and Classical Swine Fever Virus Wild Strains[J]. Acta Veterinaria et Zootechnica Sinica, 2021, 52(1): 177-184.

References 30

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Establishment and Application of a Duplex TaqMan MGB FQ-PCR for Differential Detection of African Swine Fever Virus and Classical Swine Fever Virus Wild Strains

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