Degenerate PCR method for identification of an antiapoptotic gene in BHV-1

G. Marfè, L. De Martino, G. Filomeni, C. Di Stefano, M. G. Giganti, U. Pagnini, F. Napolitano, G. Iovane, M. R. Ciriolo, P. Sinibaldi Salimei

Research output: Contribution to journalArticlepeer-review

Abstract

To investigate on the hypothetical presence of an antiapoptotic gene, we utilized the CODEHOP (COnsensus-DEgenerate Hybrid Oligonucleotide Primers) strategy amplifying unknown sequences from a background of genomic (bovine herpesvirus type-1) BHV-1 DNA. An alignment of carboxyl-terminal domains belonging to three proteins encoded by γ34.5, MyD116 and GADD34 genes, was carried out to design degenerate PCR primers in highly conserved regions. This allowed the amplification of a 110 bp fragment. This fragment was subjected to automatic sequencing and DNAsequence analysis revealed that its position resided between the nt 14363 and the nt 14438 in bovine herpesvirus type-1 (BHV-1) Cooper strain sharing an identity of 86% (UL14). Transient transfections showed that UL14 protein is efficient in protecting MDBK and K562 cells from sorbitol induced apoptosis. The protein's anti-apoptotic function may derive from its heat shock protein-like properties.

Original languageEnglish
Pages (from-to)813-823
Number of pages11
JournalJournal of Cellular Biochemistry
Volume97
Issue number4
DOIs
Publication statusPublished - Mar 1 2006

Keywords

  • Apoptosis
  • BHV-1
  • CODEHOP
  • PCR degenerate
  • UL14

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology

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