Disruption and basic functional analysis of five chromosome X novel ORFs of Saccharomyces cerevisiae reveals YJL125c as an essential gene for vegetative growth

We describe the disruption and basic functional analysis of five novel open reading frames (ORFs) discovered during the sequencing of the Saccharomyce s cerevisiae genome: YJL118w, YJL122w, YJL123c, YJL123c, YJL124c, YJL125c, located on chromosome X. Disruptions have been realized using the long-flan king homology-PCR replacement strategy (LFH-PCR; Wach ct al., 1996) in the FY1679 diploid strain. Sporulation and tetrad analysis of these heterozygou s deletants were performed, as well as a functional analysis on the haploid deleted strains: different growth conditions (complete glucose and glycero l, minimal media) at three temperatures 15, 30 and 37 degrees C were tested . Analysis revealed YJL125c as an essential gene; the four other ORFs were non-essential and showed no particular phenotype. In addition, the five kan MX4, disruption cassettes were cloned in pUG7 vector. Finally, the five ORF s with their promoter and terminator regions were cloned in the centromeric yeast vector pRS416. The vectors containing the disruption cassettes, the cognate wild-type genes, as well as the deletant yeast strains are availabl e at the EU EUROFAN (EUROSCARF, Frankfurt, DE) genetic and stock centre. Co pyright (C) 1999 John Wiley & Sons, Ltd.