Yeast vectors for integration at the HO locus.

We have constructed new yeast vectors for targeted integration of desired sequences at the Saccharomyces cerevisiae HO locus. Insertion at HO has been shown to have no effect on yeast growth, and thus these integrations should be neutral. One vector contains the KanMX selectable marker, and integrants can be selected by resistance to G418. The other vector contains the hisG-URA3-hisG cassette, and integrants can be selected by uracil prototrophy. Subsequent growth on 5-FOA permits identification of colonies where recombination between the hisG tandem repeats has led to loss of the URA3 marker and return to uracil auxotrophy. We also describe several new bacterial polylinker vectors derived from pUC21 (ampicillin resistance) and pUK21 (kanamycin resistance).

[1]  L. Hartwell,et al.  Dominant effects of tubulin overexpression in Saccharomyces cerevisiae , 1989, Molecular and cellular biology.

[2]  Nancy Kleckner,et al.  A Method for Gene Disruption That Allows Repeated Use of URA3 Selection in the Construction of Multiply Disrupted Yeast Strains , 1987, Genetics.

[3]  Vieira Jeffrey,et al.  New pUC-derived cloning vectors with different selectable markers and DNA replication origins. , 1991 .

[4]  R. Rothstein.  Targeting, disruption, replacement, and allele rescue: integrative DNA transformation in yeast. , 1991, Methods in enzymology.

[5]  L. Prakash,et al.  Yeast Saccharomyces cerevisiae selectable markers in pUC18 polylinkers , 1990, Yeast.

[6]  D. Botstein,et al.  Plasmid construction by homologous recombination in yeast. , 1987, Gene.

[7]  S. Dorland,et al.  Roles for the Saccharomyces cerevisiae SDS3, CBK1 and HYM1 genes in transcriptional repression by SIN3. , 2000, Genetics.

[8]  A. Adams,et al.  Suppressor analysis of fimbrin (Sac6p) overexpression in yeast. , 1999, Genetics.

[9]  G. Zeng.  Sticky-end PCR: new method for subcloning. , 1998, BioTechniques.

[10]  P. Philippsen,et al.  New heterologous modules for classical or PCR‐based gene disruptions in Saccharomyces cerevisiae , 1994, Yeast.

[11]  Ira Herskowitz,et al.  11 Mating-type Determination and Mating-type Interconversion in Saccharomyces cerevisiae , 1992 .

[12]  R. Akada,et al.  Screening and identification of yeast sequences that cause growth inhibition when overexpressed , 1997, Molecular and General Genetics MGG.

[13]  J. Mccusker,et al.  Three new dominant drug resistance cassettes for gene disruption in Saccharomyces cerevisiae , 1999, Yeast.

[14]  A. Bretscher,et al.  Construction of a GAL1-regulated yeast cDNA expression library and its application to the identification of genes whose overexpression causes lethality in yeast. , 1992, Genetics.

[15]  Frank Baganz,et al.  Suitability of replacement markers for functional analysis studies inSaccharomyces cerevisiae , 1997, Yeast.

[16]  J. Vieira,et al.  New pUC-derived cloning vectors with different selectable markers and DNA replication origins. , 1991, Gene.