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(New page: == Information regarding the provenance of ''Saccharomyces cerevisiae'' strain W303 == ''Kindly provided at SGD's request by Rodney Rothstein on March 10, 2005.'' The original W303 strai...)
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Revision as of 15:25, 10 May 2007

Information regarding the provenance of Saccharomyces cerevisiae strain W303

Kindly provided at SGD's request by Rodney Rothstein on March 10, 2005.

The original W303 strain is mutated in rad5-535 (an G to R change at position 535 - See Fan et al. Genetics 142:749, 1996).

  • The change is subtle resulting in a phenotype in combination with soh1 (Hannah Klein in the Fan paper), sir mutations--increased mms resistance (David Sinclair, unpublished) and no effect on recombination, UV or X-ray sensitivities (Rothstein lab, unpublished).
  • To assay for its presence in any W303 derivative strain, one can do a PCR and digest the products with MnlI, as the mutation creates a MnlI site.
    • The primers to use are:
      • 5'-gcagcaggaccatgtaaacg-3' RAD5-L
      • 5'-aaactcgttactccactgcg-3' RAD5-R
    • Run a 3% agarose gel to see the fragments.
      • In wild type: 182 bp and 155 bp.
      • In rad5-535: 155 bp, 120 bp and 62 bp.
    • The RAD5 wild type derivatives of W303 are W1588.

Some relevant information for W303:

MATa/MATalpha {leu2-3,112 trp1-1 can1-100 ura3-1 ade2-1 his3-11,15} [phi+]

  • This strain was made diploid by transforming W301-18A (Rothstein, Meth. Enzymol. 101:202-211, 1983.) with an HO-containing plasmid.
  • The diploid was dissected to obtain the isogenic MATa (W303-1A) and MATalpha (W303-1B) strains (Thomas & Rothstein, Cell 56:619-630, 1989).
  • The {brackets} in the genotype indicate that these genes are homozygous in the diploid. Each haploid strain has only a single copy the gene.
  • The [phi+] element is a non-Mendelian trait that affects the efficiency of amber suppression. Unlike the related element [psi+], this element does not affect ochre suppression.
    • ade2-1 and can1-100 are ochre-suppressible.
    • trp1-1 is amber-suppressible.
    • ura3-1 reverts at very low frequency (2 x 10e-9).
    • Both leu2-3,112 and his3-11,15 do not revert at any measurable frequency.
    • Sequence details for the relevant genes are listed in the table at the bottom of the page.

Brief description of the history of W303:

  • Many crosses were made with strains from Rothstein's Ph.D. thesis, W87 derivatives
    • see Genetics 85:35-54, 1977 and Genetics 85:55-64, 1977
    • These are mainly but not exclusively X2180-like (S288C).
  • It also got part of its genetic background from Fred Sherman's strains, D311-3A
    • see Genetics 94:871-889, 1980 and Genetics 94:891-898, 1980
  • Finally, one of the grandparents of W301-18A, D190-9C, is a real mutt, which Rothstein got from Jack Szostak and about which very little is known.

TABLE. Mutant alleles in W303.

allele nt position alteration aa change
ura3-1 701 gga > gAa Gly > Glu
trp1-1*** 247 gag > Tag Glu > amber
can1-100 139 aaa > Taa Lys > ochre
ade2-1 27** taa > ttG none
190 gaa > Taa Glu > ochre
301* aga > Gga Arg > Gly
372** gtt > gtC none
1617** acg > acA none
his3-11,15 208 G deletion -1 frameshift
319 G deletion -1 frameshift
leu2-3,112 168** gtc > gtT none
206* gtt > gCt Val > Ala
249 G insertion +1 frameshift
792 G insertion +1 frameshift
897** gtt > gtC none
898* gac > Aac Asp > Asn
  • extra mutation compared to published wild-type sequence
    ** nucleotide change compared to published wild-type sequence, but amino acid is conserved
    ***info from John McDonald, formerly of the Rothstein lab, Genetics 147:1557-1568 (1997)