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. 1996 Sep 1;24(17):3469–3471. doi: 10.1093/nar/24.17.3469

One-step PCR mediated strategy for the construction of conditionally expressed and epitope tagged yeast proteins.

D Lafontaine 1, D Tollervey 1
PMCID: PMC146098  PMID: 8811105

Abstract

With the availability of the complete yeast genomic sequence, techniques which allow the rapid functional analysis of genes of interest are of increasing importance. Here we report a technique which allows the initial characterisation of genes of interest, through the construction of conditionally expressed mutations for functional analyses and the generation of epitope-tagged fusion proteins for immuno-localisation and immuno-purification, entirely by PCR.

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Selected References

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  1. Baudin A., Ozier-Kalogeropoulos O., Denouel A., Lacroute F., Cullin C. A simple and efficient method for direct gene deletion in Saccharomyces cerevisiae. Nucleic Acids Res. 1993 Jul 11;21(14):3329–3330. doi: 10.1093/nar/21.14.3329. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Clark M. W., Yip M. L., Campbell J., Abelson J. SSB-1 of the yeast Saccharomyces cerevisiae is a nucleolar-specific, silver-binding protein that is associated with the snR10 and snR11 small nuclear RNAs. J Cell Biol. 1990 Nov;111(5 Pt 1):1741–1751. doi: 10.1083/jcb.111.5.1741. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Gietz D., St Jean A., Woods R. A., Schiestl R. H. Improved method for high efficiency transformation of intact yeast cells. Nucleic Acids Res. 1992 Mar 25;20(6):1425–1425. doi: 10.1093/nar/20.6.1425. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Lafontaine D., Vandenhaute J., Tollervey D. The 18S rRNA dimethylase Dim1p is required for pre-ribosomal RNA processing in yeast. Genes Dev. 1995 Oct 15;9(20):2470–2481. doi: 10.1101/gad.9.20.2470. [DOI] [PubMed] [Google Scholar]
  5. Längle-Rouault F., Jacobs E. A method for performing precise alterations in the yeast genome using a recycable selectable marker. Nucleic Acids Res. 1995 Aug 11;23(15):3079–3081. doi: 10.1093/nar/23.15.3079. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Schimmang T., Tollervey D., Kern H., Frank R., Hurt E. C. A yeast nucleolar protein related to mammalian fibrillarin is associated with small nucleolar RNA and is essential for viability. EMBO J. 1989 Dec 20;8(13):4015–4024. doi: 10.1002/j.1460-2075.1989.tb08584.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Sikorski R. S., Hieter P. A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae. Genetics. 1989 May;122(1):19–27. doi: 10.1093/genetics/122.1.19. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. 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]

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