Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/32836
標題: Functional analysis of virion host shutoff protein of pseudorabies virus
作者: Lin, H.W.
張天傑
Chang, Y.Y.
Wong, M.L.
Lin, J.W.
Chang, T.J.
關鍵字: pseudorabies virus
vhs gene
herpes-simplex-virus
messenger-rna degradation
dna-sequence
equine
herpesvirus-1
vhs protein
gene ul41
in-vitro
type-1
cells
identification
期刊/報告no:: Virology, Volume 324, Issue 2, Page(s) 412-418.
摘要: During lytic infection, the virion host shutoff (vhs) protein of alphaherpesviruses causes the degradation of mRNAs nonspecifically. In this work, we cloned the vhs gene (UL41 open reading frame) of pseudorabies virus (PRV; TNL strain) by PCR, and its nucleotide sequences were determined. The PCR product of vhs gene was subcloned into the prokaryotic pET32b expression vector, and production of the recombinant vhs protein was examined by SDS-PAGE. Result of Western blotting demonstrated that our recombinant vhs protein reacted with antiserum against a synthetic peptide of 17 amino acids of the vhs protein. After purification with nickel-chelate affinity chromatography, the purified recombinant vhs protein exhibited in vitro ribonuclease activity as expected. We further cloned the vhs gene into eukaryotic expression vectors and investigated the intracellular function of vhs protein by DNA transfection. By transient transfection and CAT assay, we found the CAT activity was reduced in the presence of vhs, indicating that degradation of mRNA of the CAT gene was caused by the vhs. Furthermore, our results showed that the plaque formation of pseudorabies virus was blocked by exogenous vhs. Taken together, we have cloned the vhs gene of pseudorabies virus (TNL strain) and conducted functional analysis of the recombinant vhs protein in vitro as well as in vivo. (C) 2004 Elsevier Inc. All rights reserved.
URI: http://hdl.handle.net/11455/32836
ISSN: 0042-6822
文章連結: http://dx.doi.org/10.1016/j.virol.2004.04.015
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