Establishment and Application time Quantitative PCR Assay Gene of Pseudorabies Virus in of a SYBR Green I Real- for the Detection of LAT Bama Miniature Pigs

[ Objective] This study aimed to establish a rapid, sensitive and specific SYBR Green I real-time quantitative PCR assay for the detection of latent pseudorabies virus （PRV） infection. [ Method ] SYBR Green I real-time quantitative PCR conditions and system for early detection of latent pseudorabies...

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Published in农业生物技术：英文版 Vol. 4; no. 3; pp. 62 - 66
Main Author Sufang LU Guangjun GUO Ling MO Feng WEI Bingmei DONG Wentong ZHANG Zhiqiang SHEN
Format Journal Article
LanguageEnglish
Published 2015
Subjects
LAT
SYBR
实时定量
实时荧光PCR检测
巴马小型猪
猪伪狂犬病病毒
猪繁殖与呼吸综合征病毒
病毒基因
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Summary:[ Objective] This study aimed to establish a rapid, sensitive and specific SYBR Green I real-time quantitative PCR assay for the detection of latent pseudorabies virus （PRV） infection. [ Method ] SYBR Green I real-time quantitative PCR conditions and system for early detection of latent pseudorabies virus in- fection were optimized and compared with conventional PCR to investigate the sensitivity and specificity. Subsequently, the established assay was applied to detect different clinical samples. [ Result] The sensitivity of SYBR Green I real-time quantitative PCR assay （52 copies/μl） was 1 000 times higher than that of conven- tional PCR （5.2×1^04 copies/μl） and the detection time was shortened by 1/2. The established assay could be used to detect PRV but could not be used to detect PCV2, PPV, CSFV or PRRSV. Various tissues were collected from Bama miniature pigs with latent PRV infection under sterile conditions for real-time PCR detection. Results showed that viral copy number in the brain, nasal swab, inguinal lymph node, liver, lung and spleen was above 20, while PRV was not detected in the kidney and heart tissues. [ Conclusion] The established SYBR Green I real-time quantitative PCR assay for PRV/.AT detection was specific, sensitive and rapid, which could be used for pathogen monitoring, epidemiological investigation and quantitative study of PRV.
Bibliography:SYBR Green I; Pseudorabies virus; gE deleted virus; LAT gene; Latent infection
[ Objective] This study aimed to establish a rapid, sensitive and specific SYBR Green I real-time quantitative PCR assay for the detection of latent pseudorabies virus （PRV） infection. [ Method ] SYBR Green I real-time quantitative PCR conditions and system for early detection of latent pseudorabies virus in- fection were optimized and compared with conventional PCR to investigate the sensitivity and specificity. Subsequently, the established assay was applied to detect different clinical samples. [ Result] The sensitivity of SYBR Green I real-time quantitative PCR assay （52 copies/μl） was 1 000 times higher than that of conven- tional PCR （5.2×1^04 copies/μl） and the detection time was shortened by 1/2. The established assay could be used to detect PRV but could not be used to detect PCV2, PPV, CSFV or PRRSV. Various tissues were collected from Bama miniature pigs with latent PRV infection under sterile conditions for real-time PCR detection. Results showed that viral copy number in the brain, nasal swab, inguinal lymph node, liver, lung and spleen was above 20, while PRV was not detected in the kidney and heart tissues. [ Conclusion] The established SYBR Green I real-time quantitative PCR assay for PRV/.AT detection was specific, sensitive and rapid, which could be used for pathogen monitoring, epidemiological investigation and quantitative study of PRV.
ISSN:2164-4993