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Caulimovirus Isolation and DNA Extraction

  • Simon N. Covey
  • Rob J. Noad
  • Nadia S. Al-Kaff
  • David S. Turner
Part of the Methods in Molecular Biology™ book series (MIMB, volume 81)

Abstract

Members of the caulimovirus group (1) each have a circular double-stranded DNA genome of approx 8 kbp that is encapsidated in a spherical, naked nucleocapsid of approx 50 nm diameter (Fig. 1). Caulimoviruses characteristically produce subcellular inclusion bodies in infected tissues that contain most of the virions found in cells, embedded in an apparently random manner. The host ranges of individual caulimoviruses tend to be restricted to one or a few plant families, and group members are transmitted between plants by aphid vectors. Based on possession of all, or most, of these characteristics, 12 definite, and 3 possible, members of the group have been identified (2).
Fig. 1.

Virions and genome organization of a typical caulimovirus, CaMV. The virus particles of CaMV (left) are isometric and about 50 nm in diameter (the subunits are schematic and are not a true representation). The DNA genome of CaMV is a circular double-stranded DNA of 8 kbp with three site-specific discontinuities (small closed circles). One of these (top of map) is in the DNA (−)-strand and is adjacent to the sequence homologous to the host tRNA that primes CaMV DNA synthesis by reverse transcription of 35S RNA. The other two gaps are in the (+)-strand adjacent to sequences controlling initiation of (+)-strand DNA synthesis. The genome has six major open reading frames (inner closed arrows), for which protein products have been identified. Gene I encodes a protein involved in cell-to-cell spread, gene II specifies the aphid transmission factor, the gene III product is a DNA-binding protein associated with virions, gene IV encodes the major CP, gene V specifies the viral polymerase (reverse transcriptase and RNAse H), and the gene VI product is an apparently multifunctional protein involved in transactivating viral protein synthesis, and in sequestering virions in inclusion bodies; it is also a major pathogenic determinant of symptom development. There are two major viral transcripts: 35S RNA, which probably serves two roles, one as a replication template and another as a viral mRNA; and 19S and 35S promoters (P19 and P35), respectively.

Keywords

Sodium Dodecyl Sulfate Inclusion Body Virus Yield Pancreatic Ribonuclease Sodium Sulphite 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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Copyright information

© Humana Press Inc. 1998

Authors and Affiliations

  • Simon N. Covey
    • 1
  • Rob J. Noad
    • 1
  • Nadia S. Al-Kaff
    • 1
  • David S. Turner
    • 1
  1. 1.Department of Virus ResearchJohn Innes CentreNorwichUK

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