1887

Abstract

SUMMARY

A ‘criss-cross’ DNA-RNA hybridization technique and Northern blot analysis were used to analyse late RNA transcripts in nuclear polyhedrosis virus (AcNPV)-infected cells. Infected cells were pulse-labelled with [P]orthophosphate at 24 to 30 h post-infection and cytoplasmic poly(A) RNA was isolated at 30 h post-infection. Labelled RNA was fractionated on preparative methylmercury gels and blotted under annealing conditions onto Southern blots of preparative DNA gels containing the AcNPV DNA digested with RI, HI or dIII restriction endonucleases. During hybridization the axis of electrophoresis of the DNA fragments was at right angles to the axis of electrophoresis of the RNA. The ‘criss-cross’ hybridization pattern was used to identify viral transcripts, to estimate their relative sizes and to map them on the genome. A transcription map for at least 12 late RNA species was drawn based on the consensus data obtained from several ‘criss-cross’ blot hybridizations and concurrent Northern blot analysis.

Loading

Article metrics loading...

/content/journal/jgv/10.1099/0022-1317-65-4-809
1984-04-01
2022-07-02
Loading full text...

Full text loading...

/deliver/fulltext/jgv/65/4/JV0650040809.html?itemId=/content/journal/jgv/10.1099/0022-1317-65-4-809&mimeType=html&fmt=ahah

References

  1. Adang M. J., Miller L. K. 1982; Molecular cloning of DNA complementary to mRNA of the baculovirus Autographa califomica nuclear polyhedrosis virus: location and gene products of RNA transcripts found late in infection. Journal of Virology 44:782–793
    [Google Scholar]
  2. Alwine J. C., Kemp D. J., Stark G. R. 1977; Method for detection of specific RNAs in agarose gels analyzed by diazobenzyloxymethyl-paper and hybridization with DNA probes. Proceedings of the National Academy of Sciences, U. S. A 74:5350–5354
    [Google Scholar]
  3. Bachenheimer S. L. 1980; Physical mapping of complex mRNA populations by ‘criss-cross’ DNA-RNA hybridization. Analytical Biochemistry 106:486–491
    [Google Scholar]
  4. Bailey J. M., Davidson N. 1976; Methylmercury as a reversible denaturing agent for agarose gel electrophoresis. Analytical Biochemistry 70:75–85
    [Google Scholar]
  5. Brown M., Crawford A. M., Faulkner P. 1979; Genetic analysis of a baculovirus, Autographa califomica nuclear polyhedrosis virus. I. Isolation of temperature sensitive mutants and assortment into complementation groups. Journal of Virology 31:190–198
    [Google Scholar]
  6. Brown M., Faulkner P., Cochran M. A., Chung K. L. 1980; Characterization of two morphology mutants of Autographa califomica nuclear polyhedrosis virus with large cuboidal inclusion bodies. Journal of General Virology 50:309–316
    [Google Scholar]
  7. Cochran M. A., Faulkner P. 1983; Location of homologous DNA sequences interspersed at five regions in the baculovirus AcMNPV genome. Journal of Virology 45:961–970
    [Google Scholar]
  8. Cochran M. A., Carstens E. B., Eaton B. E., Faulkner P. 1982; Molecular cloning and physical mapping of restriction endonuclease fragments of Autographa califomica nuclear polyhedrosis virus DNA. Journal of Virology 41:940–946
    [Google Scholar]
  9. Esche H., Lubbert H., Siegmann B., Doerfler W. 1982; The translational map of the Autographa califomica nuclear polyhedrosis virus (AcNPV) genome. EMBO Journal 1:1629–1633
    [Google Scholar]
  10. Kelly D. C., Lescott T. 1981; Baculovirus replication: protein synthesis mSpodoptera frugiperda cells infected with Trichoplusia ni nuclear polyhedrosis virus. Microbiologica 4:35–57
    [Google Scholar]
  11. Miller L. K., Dawes K. P. 1979; Physical map of the DNA genome of Autographa califomica nuclear polyhedrosis virus. Journal of Virology 29:1044–1055
    [Google Scholar]
  12. Rohel D. Z., Cochran M. A., Faulkner P. 1983; Characterization of two abundant mRNAs of Autographa califomica nuclear polyhedrosis virus present late in infection. Virology 124:357–365
    [Google Scholar]
  13. Smith G. E., Summers M. D. 1979; Restriction maps of five Autographa califomica MNPV variants, Trichoplusia ni MNPV and Galleria mellonella MNPV DNAs with endonucleases Smal, Kpnl, BamHl, Sad, Xhol, and,. EcoRI. Journal of Virology 30:828–838
    [Google Scholar]
  14. Smith G. E., Vlak J. M., Summers M. D. 1982; In vitro translation of Autographa califomica nuclear polyhedrosis virus early and late mRNA. Journal of Virology 44:199–208
    [Google Scholar]
  15. Smith G. E., Vlak J. M., Summers M. D. 1983; Physical analysis of Autographa califomica nuclear polyhedrosis virus transcripts for polyhedrin and 10, 000 molecular weight protein. Journal of Virology 45:215–225
    [Google Scholar]
  16. Southern E. M. 1975; Detection of specific sequences among DNA fragments separated by gel electrophoresis. Journal of Molecular Biology 98:503–517
    [Google Scholar]
  17. Vlak J. M., Smith G. E. 1982; Orientation of the genome of Autographa califomica nuclear polyhedrosis virus: a proposal. Journal of Virology 41:1118–1121
    [Google Scholar]
  18. Vlak J. M., van Der krol S. 1982; Transcription of the Autographa califomica nuclear polyhedrosis virus genome: location of late cytoplasmic mRNA. Virology 123:222–228
    [Google Scholar]
  19. Vlak J. M., Smith G. E., Summers M. D. 1981; Hybridization selection and in vitro translation of Autographa califomica nuclear polyhedrosis virus mRNA. Journal of Virology 40:762–771
    [Google Scholar]
  20. Wood H. A. 1980; Autographa califomica nuclear polyhedrosis virus-induced proteins in tissue culture. Virology 102:21–27
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/0022-1317-65-4-809
Loading
/content/journal/jgv/10.1099/0022-1317-65-4-809
Loading

Data & Media loading...

Most cited this month Most Cited RSS feed

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error