Date of Award:
5-1-2005
Document Type:
Thesis
Degree Name:
Master of Science (MS)
Department:
Biology
Committee Chair(s)
Joseph K.-K. Li
Committee
Joseph K.-K. Li
Committee
Jon Y. Takemoto
Committee
Dale L. Barnard
Committee
Stanley D. Allen
Abstract
Bluetongue virus (BTV) of the family Reoviridae has a segmented double-stranded RNA (dsRNA) genome with ten dsRNA segments which encode seven structural proteins and four nonstructural proteins. Previous studies have revealed that newly synthesized RNA transcripts during viral replication were found in equimolar ratios within BTV virions. However, synthesized mRNA transcripts intended for translation into viral proteins are not equimolar. The levels of expression of these BTV mRNAs are still unknown although the number of viral proteins present in each BTV virion had been defined. The purpose of this research was to establish a kinetic analysis of the ten BTV mRNA transcripts by a quantitative real time polymerase chain reaction (qRT-PCR) assay. Expression profiles of all ten BTV genes as well as their stability profiles for each BTV mRNA transcript were established. Concurrent with this kinetic analysis of viral mRNAs, the major BTV core protein, VP7, was characterized as a highly expressed protein by means of two-dimensional gel electrophoresis and confirmed by Western blot. We found that the levels of mRNA present were directly related to the number of viral structural proteins present in each BTV virion. These findings aid our understanding of the expression of BTV mRNA transcripts during infection in mammalian cells. These results also show that transcript levels correlated well to the expression of viral proteins and not to viral mRNA translational efficiencies. We also determined the mechanisms of BTV mRNA transcript degradation. The combined BTV profiles of mRNA and VP7 protein will improve early diagnostic testing and screening of anti-viral agents for the inhibition of BTV replication as well as provide information for gene knockouts in RNAi experiments.
Recommended Citation
Buccambuso, Margaret K., "A Kinetic Analysis of Bluetongue Virus Messenger RNA and Protein" (2005). Biology. 700.
https://digitalcommons.usu.edu/etd_biology/700
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