Skip to main content
PMC home page
Genetics logoLink to Genetics
. 1989 May;122(1):19–27. doi: 10.1093/genetics/122.1.19

A System of Shuttle Vectors and Yeast Host Strains Designed for Efficient Manipulation of DNA in Saccharomyces Cerevisiae

R S Sikorski 1, P Hieter 1
PMCID: PMC1203683  PMID: 2659436

Abstract

A series of yeast shuttle vectors and host strains has been created to allow more efficient manipulation of DNA in Saccharomyces cerevisiae. Transplacement vectors were constructed and used to derive yeast strains containing nonreverting his3, trp1, leu2 and ura3 mutations. A set of YCp and YIp vectors (pRS series) was then made based on the backbone of the multipurpose plasmid pBLUESCRIPT. These pRS vectors are all uniform in structure and differ only in the yeast selectable marker gene used (HIS3, TRP1, LEU2 and URA3). They possess all of the attributes of pBLUESCRIPT and several yeast-specific features as well. Using a pRS vector, one can perform most standard DNA manipulations in the same plasmid that is introduced into yeast.

Full Text

The Full Text of this article is available as a PDF (944.6 KB).

Selected References

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

  1. Andreadis A., Hsu Y. P., Hermodson M., Kohlhaw G., Schimmel P. Yeast LEU2. Repression of mRNA levels by leucine and primary structure of the gene product. J Biol Chem. 1984 Jul 10;259(13):8059–8062. [PubMed] [Google Scholar]
  2. Andreadis A., Hsu Y. P., Kohlhaw G. B., Schimmel P. Nucleotide sequence of yeast LEU2 shows 5'-noncoding region has sequences cognate to leucine. Cell. 1982 Dec;31(2 Pt 1):319–325. doi: 10.1016/0092-8674(82)90125-8. [DOI] [PubMed] [Google Scholar]
  3. Bajwa W., Torchia T. E., Hopper J. E. Yeast regulatory gene GAL3: carbon regulation; UASGal elements in common with GAL1, GAL2, GAL7, GAL10, GAL80, and MEL1; encoded protein strikingly similar to yeast and Escherichia coli galactokinases. Mol Cell Biol. 1988 Aug;8(8):3439–3447. doi: 10.1128/mcb.8.8.3439. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Balbás P., Soberón X., Merino E., Zurita M., Lomeli H., Valle F., Flores N., Bolivar F. Plasmid vector pBR322 and its special-purpose derivatives--a review. Gene. 1986;50(1-3):3–40. doi: 10.1016/0378-1119(86)90307-0. [DOI] [PubMed] [Google Scholar]
  5. Baldari C., Cesareni G. Plasmids pEMBLY: new single-stranded shuttle vectors for the recovery and analysis of yeast DNA sequences. Gene. 1985;35(1-2):27–32. doi: 10.1016/0378-1119(85)90154-4. [DOI] [PubMed] [Google Scholar]
  6. Benton W. D., Davis R. W. Screening lambdagt recombinant clones by hybridization to single plaques in situ. Science. 1977 Apr 8;196(4286):180–182. doi: 10.1126/science.322279. [DOI] [PubMed] [Google Scholar]
  7. Boeke J. D., LaCroute F., Fink G. R. A positive selection for mutants lacking orotidine-5'-phosphate decarboxylase activity in yeast: 5-fluoro-orotic acid resistance. Mol Gen Genet. 1984;197(2):345–346. doi: 10.1007/BF00330984. [DOI] [PubMed] [Google Scholar]
  8. Bouton A. H., Smith M. M. Fine-structure analysis of the DNA sequence requirements for autonomous replication of Saccharomyces cerevisiae plasmids. Mol Cell Biol. 1986 Jul;6(7):2354–2363. doi: 10.1128/mcb.6.7.2354. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Dobson M. J., Tuite M. F., Mellor J., Roberts N. A., King R. M., Burke D. C., Kingsman A. J., Kingsman S. M. Expression in Saccharomyces cerevisiae of human interferon-alpha directed by the TRP1 5' region. Nucleic Acids Res. 1983 Apr 25;11(8):2287–2302. doi: 10.1093/nar/11.8.2287. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Fasullo M. T., Davis R. W. Direction of chromosome rearrangements in Saccharomyces cerevisiae by use of his3 recombinational substrates. Mol Cell Biol. 1988 Oct;8(10):4370–4380. doi: 10.1128/mcb.8.10.4370. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  12. Herskowitz I. Functional inactivation of genes by dominant negative mutations. Nature. 1987 Sep 17;329(6136):219–222. doi: 10.1038/329219a0. [DOI] [PubMed] [Google Scholar]
  13. Hieter P., Pridmore D., Hegemann J. H., Thomas M., Davis R. W., Philippsen P. Functional selection and analysis of yeast centromeric DNA. Cell. 1985 Oct;42(3):913–921. doi: 10.1016/0092-8674(85)90287-9. [DOI] [PubMed] [Google Scholar]
  14. Hill J. E., Myers A. M., Koerner T. J., Tzagoloff A. Yeast/E. coli shuttle vectors with multiple unique restriction sites. Yeast. 1986 Sep;2(3):163–167. doi: 10.1002/yea.320020304. [DOI] [PubMed] [Google Scholar]
  15. Hope I. A., Struhl K. GCN4 protein, synthesized in vitro, binds HIS3 regulatory sequences: implications for general control of amino acid biosynthetic genes in yeast. Cell. 1985 Nov;43(1):177–188. doi: 10.1016/0092-8674(85)90022-4. [DOI] [PubMed] [Google Scholar]
  16. Ito H., Fukuda Y., Murata K., Kimura A. Transformation of intact yeast cells treated with alkali cations. J Bacteriol. 1983 Jan;153(1):163–168. doi: 10.1128/jb.153.1.163-168.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Johnston M., Davis R. W. Sequences that regulate the divergent GAL1-GAL10 promoter in Saccharomyces cerevisiae. Mol Cell Biol. 1984 Aug;4(8):1440–1448. doi: 10.1128/mcb.4.8.1440. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Koshland D., Hieter P. Visual assay for chromosome ploidy. Methods Enzymol. 1987;155:351–372. doi: 10.1016/0076-6879(87)55024-8. [DOI] [PubMed] [Google Scholar]
  19. Panzeri L., Philippsen P. Centromeric DNA from chromosome VI in Saccharomyces cerevisiae strains. EMBO J. 1982;1(12):1605–1611. doi: 10.1002/j.1460-2075.1982.tb01362.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Parent S. A., Fenimore C. M., Bostian K. A. Vector systems for the expression, analysis and cloning of DNA sequences in S. cerevisiae. Yeast. 1985 Dec;1(2):83–138. doi: 10.1002/yea.320010202. [DOI] [PubMed] [Google Scholar]
  21. Rose M., Grisafi P., Botstein D. Structure and function of the yeast URA3 gene: expression in Escherichia coli. Gene. 1984 Jul-Aug;29(1-2):113–124. doi: 10.1016/0378-1119(84)90172-0. [DOI] [PubMed] [Google Scholar]
  22. Rose M., Winston F. Identification of a Ty insertion within the coding sequence of the S. cerevisiae URA3 gene. Mol Gen Genet. 1984;193(3):557–560. doi: 10.1007/BF00382100. [DOI] [PubMed] [Google Scholar]
  23. Rothstein R. J. One-step gene disruption in yeast. Methods Enzymol. 1983;101:202–211. doi: 10.1016/0076-6879(83)01015-0. [DOI] [PubMed] [Google Scholar]
  24. Scherer S., Davis R. W. Replacement of chromosome segments with altered DNA sequences constructed in vitro. Proc Natl Acad Sci U S A. 1979 Oct;76(10):4951–4955. doi: 10.1073/pnas.76.10.4951. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Smith M. M., Andrésson O. S. DNA sequences of yeast H3 and H4 histone genes from two non-allelic gene sets encode identical H3 and H4 proteins. J Mol Biol. 1983 Sep 25;169(3):663–690. doi: 10.1016/s0022-2836(83)80164-8. [DOI] [PubMed] [Google Scholar]
  26. St John T. P., Scherer S., McDonell M. W., Davis R. W. Deletion analysis of the Saccharomyces GAL gene cluster. Transcription from three promoters. J Mol Biol. 1981 Oct 25;152(2):317–334. doi: 10.1016/0022-2836(81)90245-x. [DOI] [PubMed] [Google Scholar]
  27. Stinchcomb D. T., Mann C., Davis R. W. Centromeric DNA from Saccharomyces cerevisiae. J Mol Biol. 1982 Jun 25;158(2):157–190. doi: 10.1016/0022-2836(82)90427-2. [DOI] [PubMed] [Google Scholar]
  28. Struhl K., Hill D. E. Two related regulatory sequences are required for maximal induction of Saccharomyces cerevisiae his3 transcription. Mol Cell Biol. 1987 Jan;7(1):104–110. doi: 10.1128/mcb.7.1.104. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Struhl K. Nucleotide sequence and transcriptional mapping of the yeast pet56-his3-ded1 gene region. Nucleic Acids Res. 1985 Dec 9;13(23):8587–8601. doi: 10.1093/nar/13.23.8587. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Tschumper G., Carbon J. Sequence of a yeast DNA fragment containing a chromosomal replicator and the TRP1 gene. Gene. 1980 Jul;10(2):157–166. doi: 10.1016/0378-1119(80)90133-x. [DOI] [PubMed] [Google Scholar]

Articles from Genetics are provided here courtesy of Oxford University Press

RESOURCES