Research and application progresses of gE gene and its encoding gE protein in PR vaccines, diagnostic technique and epidemiological investigation are summarized, which have certain reference value for comprehensive pr...Research and application progresses of gE gene and its encoding gE protein in PR vaccines, diagnostic technique and epidemiological investigation are summarized, which have certain reference value for comprehensive prevention and control of PR and gradual purification of PR in different regions.展开更多
[ 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-t...[ 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.展开更多
In order to develop a simple and safe test for the detection of vaccinated as well as wild type Pseudorabies virus (PRV) infected pigs,the modified gE gene of PRV Ea strain,obtained by cutting the 5’ UTR using PCR an...In order to develop a simple and safe test for the detection of vaccinated as well as wild type Pseudorabies virus (PRV) infected pigs,the modified gE gene of PRV Ea strain,obtained by cutting the 5’ UTR using PCR and DNA recombinant technique,was inserted into baculovirus expression vector pFastBac 1,resulting the trans-position plamid pFE1.75.After homologous recombination,recombinant baculovirus rvBacE1.75 was gained and high level expression of glycoprotein E (gE) was observed after the infection of rvBacE1.75 to Tn-5B1-4 cells.The expression product was 80～88kD and was specific to antisera against PRV Ea strain by Western-blotting.Purified recombinant proteins were used as an antigen in Latex Agglutination Test(gE-LAT) and the test was specific,sensitive,safe and simple.展开更多
基金Supported by Natural Science Foundation of Shandong Province(ZR2014CQ012)
文摘Research and application progresses of gE gene and its encoding gE protein in PR vaccines, diagnostic technique and epidemiological investigation are summarized, which have certain reference value for comprehensive prevention and control of PR and gradual purification of PR in different regions.
基金Supported by Shandong Provincial Natural Science Foundation(ZR2012CQ012)Shandong Provincial Technical Innovation Grant(201220916006)Special Fund for Applied Technology Research and Development of Binzhou City(200706)
文摘[ 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.
文摘In order to develop a simple and safe test for the detection of vaccinated as well as wild type Pseudorabies virus (PRV) infected pigs,the modified gE gene of PRV Ea strain,obtained by cutting the 5’ UTR using PCR and DNA recombinant technique,was inserted into baculovirus expression vector pFastBac 1,resulting the trans-position plamid pFE1.75.After homologous recombination,recombinant baculovirus rvBacE1.75 was gained and high level expression of glycoprotein E (gE) was observed after the infection of rvBacE1.75 to Tn-5B1-4 cells.The expression product was 80～88kD and was specific to antisera against PRV Ea strain by Western-blotting.Purified recombinant proteins were used as an antigen in Latex Agglutination Test(gE-LAT) and the test was specific,sensitive,safe and simple.
