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. 1975 Jul;16(1):146–152. doi: 10.1128/jvi.16.1.146-152.1975

Quantitation of avian RNA tumor virus reverse transcriptase by radioimmunoassay.

A Panet, D Baltimore, T Hanafusa
PMCID: PMC354643  PMID: 48558

Abstract

A radioimmunoassay was developed that can detect and quantitate 3 ng or more of the avian RNA tumor virus reverse transcriptase. The assay detected no antigenic sites in Rous sarcoma virus alpha virions or in virions of a murine RNA tumor virus. About 70 molecules of reverse transcriptase were found per virion of avian myleloblastosis virus with this assay or with an assay based on antibody inhibition of enzymatic activity. The assay detected about 270 ng of enzyme per mg of cell protein in virus-producing cells; uninfected cells had much less antigenic material but contained some determinants able to displace radioactive antigen. No additional antigenic determinants on reverse transcriptase could be detected that were not found on the separated alpha subunit of the enzyme. Although sevenfold less sensitive than enzymatic activity as a measure of reverse transcriptase, the radioimmunoassay can detect antigen using small amounts of protein and in the presence of inhibtors.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Baltimore D. Tumor viruses: 1974. Cold Spring Harb Symp Quant Biol. 1975;39(Pt 2):1187–1200. doi: 10.1101/sqb.1974.039.01.137. [DOI] [PubMed] [Google Scholar]
  2. Bolognesi D. P., Bauer H. Polypeptides of avian RNA tumor viruses. 1. Isolation and physical and chemical analysis. Virology. 1970 Dec;42(4):1097–1112. doi: 10.1016/0042-6822(70)90357-0. [DOI] [PubMed] [Google Scholar]
  3. Chen J. H., Hanafusa H. Detection of a protein of avian leukoviruses in uninfected chick cells by radioimmunoassay. J Virol. 1974 Feb;13(2):340–346. doi: 10.1128/jvi.13.2.340-346.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Chen J. H., Hayward W. S., Hanafusa H. Avian tumor virus proteins and RNA in uninfected chicken embryo cells. J Virol. 1974 Dec;14(6):1419–1429. doi: 10.1128/jvi.14.6.1419-1429.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Fan H., Baltimore D. RNA metabolism of murine leukemia virus: detection of virus-specific RNA sequences in infected and uninfected cells and identification of virus-specific messenger RNA. J Mol Biol. 1973 Oct 15;80(1):93–117. doi: 10.1016/0022-2836(73)90235-0. [DOI] [PubMed] [Google Scholar]
  6. GREENWOOD F. C., HUNTER W. M., GLOVER J. S. THE PREPARATION OF I-131-LABELLED HUMAN GROWTH HORMONE OF HIGH SPECIFIC RADIOACTIVITY. Biochem J. 1963 Oct;89:114–123. doi: 10.1042/bj0890114. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Gibson W., Verma I. M. Studies on the reverse transcriptase of RNA tumor viruses. Structural relatedness of two subunits of avian RNA tumor viruses. Proc Natl Acad Sci U S A. 1974 Dec;71(12):4991–4994. doi: 10.1073/pnas.71.12.4991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Grandgenett D. P., Gerard G. F., Green M. A single subunit from avian myeloblastosis virus with both RNA-directed DNA polymerase and ribonuclease H activity. Proc Natl Acad Sci U S A. 1973 Jan;70(1):230–234. doi: 10.1073/pnas.70.1.230. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hanafusa H., Baltimore D., Smoler D., Watson K. F., Yaniv A., Spiegelman S. Absence of polymerase protein in virions of alpha-type rous sarcoma virus. Science. 1972 Sep 29;177(4055):1188–1191. doi: 10.1126/science.177.4055.1188. [DOI] [PubMed] [Google Scholar]
  10. Hanafusa H., Hanafusa T. Noninfectious RSV deficient in DNA polymerase. Virology. 1971 Jan;43(1):313–316. doi: 10.1016/0042-6822(71)90251-0. [DOI] [PubMed] [Google Scholar]
  11. Hurwitz J., Leis J. P. RNA-dependent DNA polymerase activity of RNA tumor viruses. I. Directing influence of DNA in the reaction. J Virol. 1972 Jan;9(1):116–129. doi: 10.1128/jvi.9.1.116-129.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kacian D. L., Watson K. F., Burny A., Spiegelman S. Purification of the DNA polymerase of avian myeloblastosis virus. Biochim Biophys Acta. 1971 Sep 24;246(3):365–383. doi: 10.1016/0005-2787(71)90773-8. [DOI] [PubMed] [Google Scholar]
  13. Kawai S., Hanafusa H. Genetic recombination with avian tumor virus. Virology. 1972 Jul;49(1):37–44. doi: 10.1016/s0042-6822(72)80005-9. [DOI] [PubMed] [Google Scholar]
  14. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  15. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  16. Moelling K. Reverse transcriptase and RNase H: present in a murine virus and in both subunits of an avian virus. Cold Spring Harb Symp Quant Biol. 1975;39(Pt 2):969–973. doi: 10.1101/sqb.1974.039.01.111. [DOI] [PubMed] [Google Scholar]
  17. Nowinski R. C., Watson K. F., Yaniv A., Spiegelman S. Serological analysis of the deoxyribonucleic acid polymerase of avian oncornaviruses. II. Comparison of avian deoxyribonucleic acid polymerases. J Virol. 1972 Nov;10(5):959–964. doi: 10.1128/jvi.10.5.959-964.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Panet A., Verma I. M., Baltimore D. Role of the subunits of the avian RNA tumor virus reverse transcriptase. Cold Spring Harb Symp Quant Biol. 1975;39(Pt 2):919–923. doi: 10.1101/sqb.1974.039.01.107. [DOI] [PubMed] [Google Scholar]
  19. Stephenson J. R., Tronick S. R., Aaronson S. A. Analysis of type specific antigenic determinants of two structural polypeptides of mouse RNA C-Type viruses. Virology. 1974 Mar;58(1):1–8. doi: 10.1016/0042-6822(74)90135-4. [DOI] [PubMed] [Google Scholar]
  20. Stephenson J. R., Wilsnack R. E., Aaronson S. A. Radioimmunoassay for avian C-type virus group-specific antigen: detection in normal and virus-transformed cells. J Virol. 1973 Jun;11(6):893–899. doi: 10.1128/jvi.11.6.893-899.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Strand M., August J. T. Structural proteins of oncogenic ribonucleic acid viruses. Interspec II, a new interspecies antigen. J Biol Chem. 1973 Aug 25;248(16):5627–5633. [PubMed] [Google Scholar]
  22. Stromberg K., Hurley N. E., Davis N. L., Rueckert R. R., Fleissner E. Structural studies of avian myeloblastosis virus: comparison of polypeptides in virion and core component by dodecyl sulfate-polyacrylamide gel electrophoresis. J Virol. 1974 Feb;13(2):513–528. doi: 10.1128/jvi.13.2.513-528.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Temin H. M., Baltimore D. RNA-directed DNA synthesis and RNA tumor viruses. Adv Virus Res. 1972;17:129–186. doi: 10.1016/s0065-3527(08)60749-6. [DOI] [PubMed] [Google Scholar]
  24. Verma I. M., Baltimore D. Purification of the RNA-directed DNA polymerase from avian myeloblastosis virus and its assay with polynucleotide templates. Methods Enzymol. 1974;29:125–130. doi: 10.1016/0076-6879(74)29015-3. [DOI] [PubMed] [Google Scholar]
  25. Vogt V. M., Eisenman R. Identification of a large polypeptide precursor of avian oncornavirus proteins. Proc Natl Acad Sci U S A. 1973 Jun;70(6):1734–1738. doi: 10.1073/pnas.70.6.1734. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Watson K. E., Nowinski R. C., Yaniv A., Spiegelman S. Serological analysis of the deoxyribonucleic acid polymerase of avian oncornaviruses. I. Preparation and characterization of monospecific antiserum with purified deoxyribonucleic acid polymerase. J Virol. 1972 Nov;10(5):951–958. doi: 10.1128/jvi.10.5.951-958.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Weissbach A., Bolden A., Muller R., Hanafusa H., Hanafusa T. Deoxyribonucleic acid polymerase activities in normal and leukovirus-infected chicken embryo cells. J Virol. 1972 Sep;10(3):321–327. doi: 10.1128/jvi.10.3.321-327.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Wright B. S., O'Brien P. A., Shibley G. P., Mayyasi S. A., Lasfargues J. C. Infection of an established mouse bone marrow cell line (JLS-V9) with Rauscher and Moloney murine leukemia viruses. Cancer Res. 1967 Sep;27(9):1672–1677. [PubMed] [Google Scholar]

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