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Validation of a reference control for an SYBR-Green fluorescence assay-based real-time PCR for detection of bovine herpesvirus 5 in experimentally exposed bovine embryos
  • Universidade Estadual Paulista (UNESP)
  • Universidade Federal de Sergipe (UFS)
  • 0890-8508
  • 1096-1194
The objective of this study was to optimize an internal control to improve SYBR-Green-based qPCR to amplify/detect the BoHV-5 US9 gene in bovine embryos produced invitro and experimentally exposed to the virus. We designed an SYBR-Green-based binding assay that is quick to perform, reliable, easily optimized and compares well with the published assay. Herein we demonstrated its general applicability to detect BoHV-5 US9 gene in bovine embryos produced invitro experimentally exposed to BoHV-5. In order to validate the assay, three different reference genes were tested; and the histone 2a gene was shown to be the most adequate for normalizing the qPCR reaction, by considering melting and standard curves ( p<0.05). On the other hand, no differences were found in the development of bovine embryos invitro whether they were exposed to BoHV-5 reference and field strains comparing to unexposed embryos. The developed qPCR assay may have important field applications as it provides an accurate BoHV-5 US9 gene detection using a proven reference gene and is considerably less expensive than the TaqMan qPCR currently employed in sanitary programs. © 2013 Elsevier Ltd.
Issue Date: 
Molecular and Cellular Probes, v. 27, n. 5-6, p. 237-242, 2013.
Time Duration: 
  • BoHV-5
  • Bovine
  • Embryos
  • Quantitative PCR
  • Bovine herpes virus
  • computer program
  • controlled study
  • cow
  • embryo
  • gene
  • histone 2a gene
  • nonhuman
  • oocyte
  • priority journal
  • real time polymerase chain reaction
  • sybr green fluorescence assay
  • zygote
  • Bovinae
  • Bovine herpesvirus 5
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Appears in Collections:Artigos, TCCs, Teses e Dissertações da Unesp

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