Skip to main content
PMC home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1982 Dec;79(23):7415–7419. doi: 10.1073/pnas.79.23.7415

Vaccinia virus: a selectable eukaryotic cloning and expression vector.

M Mackett, G L Smith, B Moss
PMCID: PMC347350  PMID: 6296831

Abstract

Foreign DNA was inserted into two nonessential regions of the vaccinia virus genome by homologous recombination in cells infected with virus and transfected with plasmids containing the foreign DNA elements flanked by vaccinia virus DNA. Thymidine kinase-negative (TK-) recombinants were selected after inserting foreign DNA into the coding region of the TK gene of wild-type vaccinia virus; TK+ recombinants were selected after inserting the herpesvirus TK gene into TK- mutants of vaccinia virus. For TK+ expression, it was necessary to insert a 275-base-pair DNA fragment containing the initiation site and sequences upstream of an early vaccinia virus transcript next to the coding sequences of the herpesvirus gene. The unique ability of the herpesvirus TK to phosphorylate 125I-labeled deoxycytidine provided independent confirmation of gene expression. These studies demonstrate the use of vaccinia virus as a selectable cloning and expression vector, confirm the map location of the vaccinia virus TK gene, and provide initial information regarding the location of vaccinia virus transcriptional regulatory sequences.

Full text

PDF

Images in this article

7416Image
on p.7416
7418Image
on p.7418
7418Image
on p.7418
7418Image
on p.7418
7418Image
on p.7418
7418Image
on p.7418
7418Image
on p.7418
7418Image
on p.7418

Selected References

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

  1. Birnboim H. C., Doly J. A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res. 1979 Nov 24;7(6):1513–1523. doi: 10.1093/nar/7.6.1513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bolivar F., Rodriguez R. L., Greene P. J., Betlach M. C., Heyneker H. L., Boyer H. W., Crosa J. H., Falkow S. Construction and characterization of new cloning vehicles. II. A multipurpose cloning system. Gene. 1977;2(2):95–113. [PubMed] [Google Scholar]
  3. Camiener G. W. Studies of the enzymatic deamination of ara-cytidine. V. inhibition in vitro and in vivo by tetrahydrouridine and other reduced pyrimidine nucleosides. Biochem Pharmacol. 1968 Sep;17(9):1981–1991. doi: 10.1016/0006-2952(68)90114-7. [DOI] [PubMed] [Google Scholar]
  4. Cooper J. A., Wittek R., Moss B. Hybridization selection and cell-free translation of mRNA's encoded within the inverted terminal repetition of the vaccinia virus genome. J Virol. 1981 Jan;37(1):284–294. doi: 10.1128/jvi.37.1.284-294.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Dales S., Pogo B. G. Biology of poxviruses. Virol Monogr. 1981;18:1–109. doi: 10.1007/978-3-7091-8625-1. [DOI] [PubMed] [Google Scholar]
  6. Denhardt D. T. A membrane-filter technique for the detection of complementary DNA. Biochem Biophys Res Commun. 1966 Jun 13;23(5):641–646. doi: 10.1016/0006-291x(66)90447-5. [DOI] [PubMed] [Google Scholar]
  7. Enquist L. W., Vande Woude G. F., Wagner M., Smiley J. R., Summers W. C. Construction and characterization of a recombinant plasmid encoding the gene for the thymidine kinase of Herpes simplex type 1 virus. Gene. 1979 Nov;7(3-4):335–342. doi: 10.1016/0378-1119(79)90052-0. [DOI] [PubMed] [Google Scholar]
  8. Gruss P., Khoury G. Expression of simian virus 40-rat preproinsulin recombinants in monkey kidney cells: use of preproinsulin RNA processing signals. Proc Natl Acad Sci U S A. 1981 Jan;78(1):133–137. doi: 10.1073/pnas.78.1.133. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hamer D. H., Leder P. Expression of the chromosomal mouse Beta maj-globin gene cloned in SV40. Nature. 1979 Sep 6;281(5726):35–40. doi: 10.1038/281035a0. [DOI] [PubMed] [Google Scholar]
  10. McKnight S. L. The nucleotide sequence and transcript map of the herpes simplex virus thymidine kinase gene. Nucleic Acids Res. 1980 Dec 20;8(24):5949–5964. doi: 10.1093/nar/8.24.5949. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Moss B., Winters E., Cooper J. A. Deletion of a 9,000-base-pair segment of the vaccinia virus genome that encodes nonessential polypeptides. J Virol. 1981 Nov;40(2):387–395. doi: 10.1128/jvi.40.2.387-395.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Moss B., Winters E., Cooper N. Instability and reiteration of DNA sequences within the vaccinia virus genome. Proc Natl Acad Sci U S A. 1981 Mar;78(3):1614–1618. doi: 10.1073/pnas.78.3.1614. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Moyer R. W., Rothe C. T. The white pock mutants of rabbit poxvirus. I. Spontaneous host range mutants contain deletions. Virology. 1980 Apr 15;102(1):119–132. doi: 10.1016/0042-6822(80)90075-6. [DOI] [PubMed] [Google Scholar]
  14. Mulligan R. C., Howard B. H., Berg P. Synthesis of rabbit beta-globin in cultured monkey kidney cells following infection with a SV40 beta-globin recombinant genome. Nature. 1979 Jan 11;277(5692):108–114. doi: 10.1038/277108a0. [DOI] [PubMed] [Google Scholar]
  15. Nakano E., Panicali D., Paoletti E. Molecular genetics of vaccinia virus: demonstration of marker rescue. Proc Natl Acad Sci U S A. 1982 Mar;79(5):1593–1596. doi: 10.1073/pnas.79.5.1593. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Panicali D., Davis S. W., Mercer S. R., Paoletti E. Two major DNA variants present in serially propagated stocks of the WR strain of vaccinia virus. J Virol. 1981 Mar;37(3):1000–1010. doi: 10.1128/jvi.37.3.1000-1010.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Rhim J. S., Cho H. Y., Huebner R. J. Non-producer human cells induced by murine sarcoma virus. Int J Cancer. 1975 Jan 15;15(1):23–29. doi: 10.1002/ijc.2910150104. [DOI] [PubMed] [Google Scholar]
  18. Rigby P. W., Dieckmann M., Rhodes C., Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. doi: 10.1016/0022-2836(77)90052-3. [DOI] [PubMed] [Google Scholar]
  19. SZYBALSKA E. H., SZYBALSKI W. Genetics of human cess line. IV. DNA-mediated heritable transformation of a biochemical trait. Proc Natl Acad Sci U S A. 1962 Dec 15;48:2026–2034. doi: 10.1073/pnas.48.12.2026. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Sarver N., Gruss P., Law M. F., Khoury G., Howley P. M. Bovine papilloma virus deoxyribonucleic acid: a novel eucaryotic cloning vector. Mol Cell Biol. 1981 Jun;1(6):486–496. doi: 10.1128/mcb.1.6.486. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Shimotohno K., Temin H. M. Formation of infectious progeny virus after insertion of herpes simplex thymidine kinase gene into DNA of an avian retrovirus. Cell. 1981 Oct;26(1 Pt 1):67–77. doi: 10.1016/0092-8674(81)90034-9. [DOI] [PubMed] [Google Scholar]
  22. Smiley J. R., Wagner M. J., Summers W. P., Summers W. C. Genetic and physical evidence for the polarity of transcription of the thymidine kinase gene of herpes simplex virus. Virology. 1980 Apr 15;102(1):83–93. doi: 10.1016/0042-6822(80)90072-0. [DOI] [PubMed] [Google Scholar]
  23. Solnick D. Construction of an adenovirus-SV40 recombinant producing SV40 T antigen from an adenovirus late promoter. Cell. 1981 Apr;24(1):135–143. doi: 10.1016/0092-8674(81)90509-2. [DOI] [PubMed] [Google Scholar]
  24. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  25. Summers W. C., Summers W. P. [125I]deoxycytidine used in a rapid, sensitive, and specific assay for herpes simplex virus type 1 thymidine kinase. J Virol. 1977 Oct;24(1):314–318. doi: 10.1128/jvi.24.1.314-318.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Thummel C., Tjian R., Grodzicker T. Expression of SV40 T antigen under control of adenovirus promoters. Cell. 1981 Mar;23(3):825–836. doi: 10.1016/0092-8674(81)90447-5. [DOI] [PubMed] [Google Scholar]
  27. Venkatesan S., Baroudy B. M., Moss B. Distinctive nucleotide sequences adjacent to multiple initiation and termination sites of an early vaccinia virus gene. Cell. 1981 Sep;25(3):805–813. doi: 10.1016/0092-8674(81)90188-4. [DOI] [PubMed] [Google Scholar]
  28. Vogelstein B., Gillespie D. Preparative and analytical purification of DNA from agarose. Proc Natl Acad Sci U S A. 1979 Feb;76(2):615–619. doi: 10.1073/pnas.76.2.615. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Wagner M. J., Sharp J. A., Summers W. C. Nucleotide sequence of the thymidine kinase gene of herpes simplex virus type 1. Proc Natl Acad Sci U S A. 1981 Mar;78(3):1441–1445. doi: 10.1073/pnas.78.3.1441. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Wahl G. M., Stern M., Stark G. R. Efficient transfer of large DNA fragments from agarose gels to diazobenzyloxymethyl-paper and rapid hybridization by using dextran sulfate. Proc Natl Acad Sci U S A. 1979 Aug;76(8):3683–3687. doi: 10.1073/pnas.76.8.3683. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Wei C. M., Gibson M., Spear P. G., Scolnick E. M. Construction and isolation of a transmissible retrovirus containing the src gene of Harvey murine sarcoma virus and the thymidine kinase gene of herpes simplex virus type 1. J Virol. 1981 Sep;39(3):935–944. doi: 10.1128/jvi.39.3.935-944.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Weir J. P., Bajszár G., Moss B. Mapping of the vaccinia virus thymidine kinase gene by marker rescue and by cell-free translation of selected mRNA. Proc Natl Acad Sci U S A. 1982 Feb;79(4):1210–1214. doi: 10.1073/pnas.79.4.1210. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Winberg G., Hammarskjöld M. L. Isolation of DNA from agarose gels using DEAE-paper. Application to restriction site mapping of adenovirus type 16 DNA. Nucleic Acids Res. 1980 Jan 25;8(2):253–264. doi: 10.1093/nar/8.2.253. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Wittek R., Cooper J. A., Barbosa E., Moss B. Expression of the vaccinia virus genome: analysis and mapping of mRNAs encoded within the inverted terminal repetition. Cell. 1980 Sep;21(2):487–493. doi: 10.1016/0092-8674(80)90485-7. [DOI] [PubMed] [Google Scholar]
  35. Wittek R., Cooper J. A., Moss B. Transcriptional and translational mapping of a 6.6-kilobase-pair DNA fragment containing the junction of the terminal repetition and unique sequence at the left end of the vaccinia virus genome. J Virol. 1981 Sep;39(3):722–732. doi: 10.1128/jvi.39.3.722-732.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Wittek R., Moss B. Colinearity of RNAs with the vaccinia virus genome: anomalies with two complementary early and late RNAs result from a small deletion or rearrangement within the inverted terminal repetition. J Virol. 1982 May;42(2):447–455. doi: 10.1128/jvi.42.2.447-455.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES