Three new dominant drug resistance cassettes for gene disruption in Saccharomyces cerevisiae

Disruption‐deletion cassettes are powerful tools used to study gene function in many organisms, including Saccharomyces cerevisiae. Perhaps the most widely useful of these are the heterologous dominant drug resistance cassettes, which use antibiotic resistance genes from bacteria and fungi as selectable markers. We have created three new dominant drug resistance cassettes by replacing the kanamycin resistance (kanr) open reading frame from the kanMX3 and kanMX4 disruption‐deletion cassettes (Wach et al., 1994) with open reading frames conferring resistance to the antibiotics hygromycin B (hph), nourseothricin (nat) and bialaphos (pat). The new cassettes, pAG25 (natMX4), pAG29 (patMX4), pAG31 (patMX3), pAG32 (hphMX4), pAG34 (hphMX3) and pAG35 (natMX3), are cloned into pFA6, and so are in all other respects identical to pFA6–kanMX3 and pFA6–kanMX4. Most tools and techniques used with the kanMX plasmids can also be used with the hph, nat and patMX containing plasmids. These new heterologous dominant drug resistance cassettes have unique antibiotic resistance phenotypes and do not affect growth when inserted into the ho locus. These attributes make the cassettes ideally suited for creating S. cerevisiae strains with multiple mutations within a single strain. Copyright © 1999 John Wiley & Sons, Ltd.

[1]  Xuewen Pan,et al.  Heterologous URA3MX cassettes for gene replacement in Saccharomyces cerevisiae , 1999, Yeast.

[2]  J. Boeke,et al.  Designer deletion strains derived from Saccharomyces cerevisiae S288C: A useful set of strains and plasmids for PCR‐mediated gene disruption and other applications , 1998, Yeast.

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

[4]  L. W. Parks,et al.  Assessment of the essentiality of ERG genes late in ergosterol biosynthesis in Saccharomyces cerevisiae , 1997, Current Genetics.

[5]  M. Berg,et al.  Expression Cassettes for Formaldehyde and Fluoroacetate Resistance, Two Dominant Markers in Saccharomyces cerevisiae , 1997 .

[6]  D Botstein,et al.  Functional Analysis of the Genes of Yeast Chromosome V by Genetic Footprinting , 1996, Science.

[7]  Ronald W. Davis,et al.  Quantitative phenotypic analysis of yeast deletion mutants using a highly parallel molecular bar–coding strategy , 1996, Nature Genetics.

[8]  J. Hegemann,et al.  Green fluorescent protein as a marker for gene expression and subcellular localization in budding yeast , 1996, Yeast.

[9]  D. Botstein,et al.  Functional analysis reports. Precise gene disruption in Saccharomyces cerevisiae by double fusion polymerase chain reaction , 1995, Yeast.

[10]  E. Jacobs,et al.  A method for performing precise alterations in the yeast genome using a recycable selectable marker. , 1995, Nucleic acids research.

[11]  D Botstein,et al.  Genetic footprinting: a genomic strategy for determining a gene's function given its sequence. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[12]  A. Willems,et al.  Studies on the transformation of intact yeast cells by the LiAc/SS‐DNA/PEG procedure , 1995, Yeast.

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

[14]  R. Mortimer,et al.  Genome renewal: A new phenomenon revealed from a genetic study of 43 strains of Saccharomyces cerevisiae derived from natural fermentation of grape musts , 1994, Yeast.

[15]  Ronald W. Davis,et al.  Genetic characterization of pathogenic Saccharomyces cerevisiae isolates. , 1994, Genetics.

[16]  G. B. Wisdom,et al.  Current protocols in immunology , 1993 .

[17]  C. Smith,et al.  Sequence and transcriptional analysis of the nourseothricin acetyltransferase-encoding gene nat1 from Streptomyces noursei. , 1993, Gene.

[18]  M. Kapoor,et al.  An electroporation-based system for high-efficiency transformation of germinated conidia of filamentous fungi. , 1991, Canadian journal of microbiology.

[19]  R. Hall,et al.  Optimized vectors and selection for transformation of Neurospora crassa and Aspergillus nidulans to bleomycin and phleomycin resistance. , 1990, Gene.

[20]  C. Rosen,et al.  A rapid method for the construction of synthetic genes using the polymerase chain reaction. , 1990, BioTechniques.

[21]  M. Vidal,et al.  The yeast gene ERG6 is required for normal membrane function but is not essential for biosynthesis of the cell-cycle-sparking sterol , 1989, Molecular and cellular biology.

[22]  R. Sikorski,et al.  A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae. , 1989, Genetics.

[23]  S. Ho,et al.  Site-directed mutagenesis by overlap extension using the polymerase chain reaction. , 1989, Gene.

[24]  A. Pühler,et al.  Nucleotide sequence of the phosphinothricin N-acetyltransferase gene from Streptomyces viridochromogenes Tü494 and its expression in Nicotiana tabacum. , 1988, Gene.

[25]  A. Pühler,et al.  Cloning of a phosphinothricin N-acetyltransferase gene from Streptomyces viridochromogenes Tü494 and its expression in Streptomyces lividans and Escherichia coli. , 1988, Gene.

[26]  P. Carter,et al.  Site-directed mutagenesis. , 1986, The Biochemical journal.

[27]  T. Platt.  Toxicity of 2-deoxygalactose to Saccharomyces cerevisiae cells constitutively synthesizing galactose-metabolizing enzymes , 1984, Molecular and cellular biology.

[28]  Z. Olempska-Beer,et al.  Initiation of meiosis and sporulation of Saccharomyces cerevisiae by sulfur or guanine deprivation. , 1984, Developmental biology.

[29]  J. Ramos,et al.  The action of 2-amino-4-(methylphosphinyl)-butanoic acid (phosphinothricin) and its 2-oxo-derivative on the metabolism of cyanobacteria and higher plants , 1984 .

[30]  J. Davies,et al.  Plasmid-encoded hygromycin B resistance: the sequence of hygromycin B phosphotransferase gene and its expression in Escherichia coli and Saccharomyces cerevisiae. , 1983, Gene.

[31]  T. Cooper.  Transport in Saccharomyces cerevisiae , 1982 .

[32]  M. Breitenbach,et al.  Metabolism of myo-inositol during sporulation of myo-inositol-requiring Saccharomyces cerevisiae , 1981, Journal of bacteriology.

[33]  S. Henry,et al.  Yeast mutants auxotrophic for choline or ethanolamine , 1980, Journal of bacteriology.

[34]  T. Manney,et al.  The CAN1 locus of Saccharomyces cerevisiae: fine-structure analysis and forward mutation rates. , 1979, Genetics.

[35]  G. Fink,et al.  Methods in yeast genetics , 1979 .

[36]  W. Daily,et al.  Hygromycin. I. Preliminary studies on the production and biologic activity of a new antibiotic. , 1953, Antibiotics & chemotherapy.

[37]  P. Burkholder,et al.  Studies on the Biochemical Genetics of Yeast. , 1949, Proceedings of the National Academy of Sciences of the United States of America.