Three new dominant drug resistance cassettes for gene disruption inSaccharomyces cerevisiae

TLDR

Disruption‑deletion cassettes are powerful tools for studying gene function in many organisms, including Saccharomyces cerevisiae, and the most widely useful of these are heterologous dominant drug resistance cassettes that use antibiotic resistance genes from bacteria and fungi as selectable markers. The authors created three new dominant drug resistance cassettes by replacing the kanamycin resistance ORF in kanMX3 and kanMX4 with ORFs conferring resistance to hygromycin B, nourseothricin, and bialaphos. The new cassettes (pAG25, pAG29, pAG31, pAG32, pAG34, pAG35) were cloned into.

Abstract

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.