Analysis of genetic interactions on a genome-wide scale in budding yeast: Diploid-based synthetic lethality analysis by microarray

Pamela B. Meluh, Xuewen Pan, Daniel S. Yuan, Carol Tiffany, Ou Chen, Sharon Sookhai-Mahadeo, Xiaoling Wang, Brian D. Peyser, Rafael Irizarry, Forrest A. Spencer, Jef D. Boeke

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

Comprehensive collections of open reading frame (ORF) deletion mutant strains exist, for the budding yeast Saccharomyces cerevisiae. With great prescience, these strains were designed with short molecular bar codes or TAGs that uniquely mark each deletion allele, flanked by shared priming sequences. These features have enabled researchers to handle yeast mutant collections as complex pools of -6000 strains. The presence of any individual mutant within a pool can be assessed indirectly by measuring the relative abundance of its corresponding TAG(s) in genomic DNA prepared from the pool. This is readily accomplished by wholesale polymerase chain reaction (PCR) amplification of the TAGs using fluorescent oligonucleotide primers that recognize the common flanking sequences, followed by hybridization of the labeled PCR products to a TAG oligonucleotide microarray. Here we describe a method-diploid-based synthetic lethality analysis by microarray (dSLAM)-whereby such pools can be manipulated to rapidly construct and assess the fitness of 6000 double-mutant strains in a single experiment. Analysis of doublemutant strains is of growing importance in defining the spectrum of essential cellular functionalities and in understanding how these functionalities interrelate.

Original languageEnglish (US)
Title of host publicationMicrobial Gene Essentiality Protocols and Bioinformatics
Pages221-247
Number of pages27
StatePublished - Dec 1 2007

Publication series

NameMethods in Molecular Biology
Volume416
ISSN (Print)1064-3745

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Keywords

  • Genetic interaction
  • Molecular barcode
  • Oligonucleotide microarray
  • SLAM
  • Synthetic lethality
  • Yeast knock-out strains

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

Cite this

Meluh, P. B., Pan, X., Yuan, D. S., Tiffany, C., Chen, O., Sookhai-Mahadeo, S., Wang, X., Peyser, B. D., Irizarry, R., Spencer, F. A., & Boeke, J. D. (2007). Analysis of genetic interactions on a genome-wide scale in budding yeast: Diploid-based synthetic lethality analysis by microarray. In Microbial Gene Essentiality Protocols and Bioinformatics (pp. 221-247). (Methods in Molecular Biology; Vol. 416).