The short flanking homology PCR strategy (Wach et al., 1994) was used to disrupt six open reading frames (ORFs) on chromosome X of diploid strains (FY1679 and W303) of the yeast Saccharomyces cerevisiae. Two of the six ORFs analysed (YJL069c and YJL066c) display no similarity to known sequences. Three others (YJL065c, YJL068c, and YJL070c) are similar to those respectively encoding the DNA polymerase epsilon subunit c, human esterase D and rat AMP deaminase 1. YJL071w has recently been identified as the ARG2 gene coding for acetylglutamate synthase. Inactivation of the YJL069c gene proved lethal and the yjl071w haploid disruptants were auxotrophic for arginine. For the four other gene inactivations, neither the heterozygous deletion diploids nor the corresponding haploid deletion mutants displayed any special phenotype when grown on rich glycerol or glucose medium or on synthetic minimal medium at three different temperatures, or on media containing compounds interfering with nucleic acid or protein synthesis. Mating and sporulation efficiencies were the same for the viable disruptants as for wild-type cells. The six kanMX4 disruption cassettes were cloned into the pUG7 vector and each of the cognate wild-type genes was inserted into the pRS416 centromeric plasmid. All strains and plasmids have been deposited in the EUROFAN collection (EUROSCARF, K. -D. Entian, Frankfurt, Germany).

• PMID: 10509023
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Journal Article | Research Support, Non-U.S. Gov't

Authors

Vandenbol M, Portetelle D

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