Amplification, cloning, sequencing, and expression of the putative gene encoding the major envelope protein of a noncytopathic strain of bovine viral diarrhea virus

Author

Melissa A. Kennedy

Date of Award

12-1991

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major

Comparative and Experimental Medicine

Major Professor

Leon N.D. Potgieter

Committee Members

Michael Breider, David Brian, Al Ichiki

Abstract

An investigation was undertaken to clone, analyze, and express the gene encoding the major envelope protein of Bovine Viral Diarrhea Virus. Polymerase chain reaction was used to generate cDNA encoding the major envelope protein of noncytopathic BVDV strain 2724 for cloning. Primers for cDNA synthesis and amplification were based on published sequence data. The 5' ends of the primers were modified by the addition of restriction sites to allow for ligation into a plasmid vector. Recombinant plasmids were sequenced by dideoxy chain termination. The insert was 1323 nucleotides in length capable of encoding a protein of 441 amino acids. Sequence analysis furthered the characterization of this genomic region of BVDV. Comparisons of this region to flaviviruses and to other pestiviruses revealed a high degree of homology between BVDV strain 2724 and the pestiviruses, but not with the flaviviruses. This construct was expressed in a cell-free translation system and data suggested that this protein was translocated and glycosylated. Results from the immunoprecipitation of the translation product indicated it was an authentic BVDV-specific protein. Identification of the putative transmembrane domain was also accomplished. It was concluded that the putative major envelope protein of BVDV strain 2724 is highly homologous to the major envelope proteins of the pestiviruses whose genomes have been cloned and sequenced. Additionally, this protein of BVDV strain 2724 was determined to be translocated, glycosylated, and anchored. The results of this study may have implications for the control and prevention of BVDV infection through improved diagnostics and immunoprophylaxis.

Recommended Citation

Kennedy, Melissa A., "Amplification, cloning, sequencing, and expression of the putative gene encoding the major envelope protein of a noncytopathic strain of bovine viral diarrhea virus. " PhD diss., University of Tennessee, 1991.
https://trace.tennessee.edu/utk_graddiss/11144