Functional analysis of six genes from chromosomes XIV and XV of Saccharomyces cerevisiae reveals YOR145c as an essential gene and YNL059c/ARP5 as a strain‐dependent essential gene encoding nuclear proteins

• Published in: Yeast (Chichester, England) Vol. 16; no. 11; pp. 1025 - 1033
• Main Authors: Grava, Sandrine, Dumoulin, Pascal, Madania, Ammar, Tarassov, Ivan, Winsor, Barbara
• Format: Journal Article
• Language: English
• Published: Chichester, UK John Wiley & Sons, Ltd 01.08.2000; Wiley
• Subjects: Actins; Actins - metabolism; ARP5 gene; Cellular Biology; Chromosomal Proteins, Non-Histone - genetics; Chromosomal Proteins, Non-Histone - metabolism; chromosome XIV; chromosome XV; Chromosomes, Fungal; Chromosomes, Fungal - genetics; Cloning, Molecular; DNA Repair; Fungal Proteins; Gene Deletion; Genes, Essential; Green Fluorescent Proteins; Life Sciences; Luminescent Proteins; Luminescent Proteins - genetics; Luminescent Proteins - metabolism; Nuclear Proteins; Nuclear Proteins - genetics; Nuclear Proteins - metabolism; Open Reading Frames; Open Reading Frames - genetics; Polymerase Chain Reaction; Saccharomyces cerevisiae; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae - growth & development; Saccharomyces cerevisiae - metabolism; Saccharomyces cerevisiae Proteins - genetics; Saccharomyces cerevisiae Proteins - metabolism; Sequence Analysis, DNA; Species Specificity; Tubulin; Tubulin - metabolism; Y0R145c gene; YNL059c gene; YOR145c; YOR152c; YOR161c; YOR165w
• ISSN: 0749-503X; 1097-0061
• DOI: 10.1002/1097-0061(200008)16:11<1025::AID-YEA602>3.0.CO;2-1

We report here basic functional analysis of strains deleted for six open reading frames (ORFs), YNL059c and YNL148c from chromosome XIV and YOR145c, YOR152c, YOR161c and YOR162c from chromosome XV of Saccharomyces cerevisiae. ORFs were replaced with the KanMX4 resistance marker using a long flanking homology PCR strategy in FY1679 and W303 diploid strains. Replacement cassettes were constructed in plasmid pUG7 and the cognate wild‐type genes were recovered by gap repair. Sporulation and tetrad analysis revealed that deletion of YNL059c/ARP5 was lethal for vegetative growth in strain W303 and caused severe growth defects in strain FY1679 while YOR145c was essential for growth in both strains. Fusion of the green fluorescent protein (GFP) gene to the 3′ ends of the YNL059c/ARP5 and YOR145c coding sequences created functional chimeric genes at the respective chromosomal loci. Both Arp5–GFP and Yor145–GFP localized to the nucleus, Yor145–GFP concentrating in the nucleolus. The vectors containing the deletion cassettes and the cognate wild‐type genes, the oligonucleotides, and the deletant strains are available from the EUROFAN resource centre EUROSCARF (Frankfurt). Copyright © 2000 John Wiley & Sons, Ltd.