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Proceedings of the National Academy of Sciences of the United States of America1981; 78(11); 6684-6688; doi: 10.1073/pnas.78.11.6684

Molecular cloning of equine herpesvirus type 1 DNA: analysis of standard and defective viral genomes and viral sequences in oncogenically transformed cells.

Abstract: Genomic DNA sequences of equine herpesvirus type 1 (EHV-1) have been cloned as BamHI and EcoRI restriction fragments into the plasmid pBR322 and propagated in Escherichia coli. With the exception of two EcoRI restriction fragments that reside in the S region of the viral genome, all of the cloned fragments demonstrated the same electrophoretic mobilities, restriction cleavage sites, and blot-hybridization patterns as did the parent fragments produced by BamHI or EcoRI digestion of virion DNA. The EcoRI J fragment and the BamHI E fragment of the L-region terminus were cloned after the addition of appropriate linker oligonucleotides. Fragments originating from each of the two isomeric forms of EHV-1 DNA were contained in this library of clones. Supramolar DNA fragments present only in the DNA of defective interfering (DI) particles of EHV-1 were generated from Bgl II digestion of DNA preparations enriched for EHV-1 DI particles and were cloned as Bgl II and EcoRI fragments into the plasmid vector. The cloned viral sequences represented in this defective genome mapped to the S region of EHV-1 DNA. Blot-hybridization analysis of EHV-1 transformed and tumor cell DNAs with the cloned BamHI B fragment confirmed that subgenomic viral sequences are present and indicated that those sequences map to the viral genome between 0.32 and 0.43 map unit.
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Publication Date: 1981-11-01 PubMed ID: 6273883PubMed Central: PMC349114DOI: 10.1073/pnas.78.11.6684Google Scholar: Lookup
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  • Journal Article
  • Research Support
  • Non-U.S. Gov't
  • Research Support
  • U.S. Gov't
  • Non-P.H.S.
  • Research Support
  • U.S. Gov't
  • P.H.S.

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

This research investigates the DNA structure of equine herpesvirus type 1 (EHV-1). The DNA of the virus was inserted into a plasmid and then copied or cloned in E.coli to better understand their genomic composition. This study also looked at defective interfering particles of the virus and found conclusions about how these might interact with regular virus particles.

Cloning of EHV-1 DNA

  • The DNA of equine herpesvirus type 1 (EHV-1) was cloned using special enzymes called BamHI and EcoRI which allow specific sections of the DNA to be cut out and inserted into a plasmid in Escherichia coli (a type of bacteria).
  • In the plasmid, the DNA sequence can generate copies of itself which allows further analysis.
  • Most of the cloned fragments showed the same traits as the original EHV-1 DNA fragments, the exceptions were two fragments that are part of the ‘S region’ of the viral genome.

Linker Oligonucleotides and Defective Interfering Particles

  • Two parts of the virus’s DNA, the EcoRI J fragment and the BamHI E fragment, were specifically cloned after adding linker oligonucleotides, short synthetic DNA sequences that help in the cloning process.
  • Defective interfering (DI) particles of EHV-1 which are incomplete virus particles that can interfere with the replication of proper virus particles, were also studied. DNA was prepared from a population of DI particles and cloned into the plasmid.
  • The cloned DI sequences matched with the S region of the EHV-1 DNA, indicating where these defective particles fit into the picture of the entire EHV-1 genome.

Confirmation of Viral Sequences in Transformed and Tumor Cells

  • The presence of specific virus sequences was confirmed within the DNA of cells that had been transformed by EHV-1. This could indicate that the virus has altered the DNA of these cells to facilitate its own reproduction.
  • The sequences present were found to be between the 0.32 and 0.43 map unit of the viral genome, hence this region may play a key role in the virus’s oncogenic (cancer-causing) activity.

Cite This Article

APA
Robinson RA, Tucker PW, Dauenhauer SA, O'Callaghan DJ. (1981). Molecular cloning of equine herpesvirus type 1 DNA: analysis of standard and defective viral genomes and viral sequences in oncogenically transformed cells. Proc Natl Acad Sci U S A, 78(11), 6684-6688. https://doi.org/10.1073/pnas.78.11.6684

Publication

Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424
NlmUniqueID: 7505876
Country: United States
Language: English
Volume: 78
Issue: 11
Pages: 6684-6688

Researcher Affiliations

Robinson, R A
    Tucker, P W
      Dauenhauer, S A
        O'Callaghan, D J

          MeSH Terms

          • Animals
          • Cell Line
          • Cell Transformation, Neoplastic
          • Cell Transformation, Viral
          • Cloning, Molecular
          • Cricetinae
          • DNA Restriction Enzymes
          • DNA, Recombinant / metabolism
          • DNA, Viral / genetics
          • Embryo, Mammalian
          • Escherichia coli / genetics
          • Herpesviridae / genetics
          • Nucleic Acid Hybridization
          • Plasmids

          Grant Funding

          • AI-02032 S-507-RR05386 / NIAID NIH HHS

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          Citations

          This article has been cited 12 times.
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