Genome-Wide Transcriptome Analysis in Yeast Using High-Density Tiling Arrays

Lior David, Sandra Clauder-Münster, Lars M. Steinmetz*

*Corresponding author for this work

Research output: Chapter in Book/Report/Conference proceedingChapterpeer-review

7 Scopus citations

Abstract

In the last decade, it became clear that transcription goes far beyond that of protein-coding genes. Most RNA molecules are transcribed from intergenic regions or introns and exhibit much variability in size, expression level, secondary structure, and evolutionary conservation. While for several types of non-coding RNAs some cellular functions have been reported, like for micro-RNAs and small nucleolar RNAs, for most others no indications of function or regulation have so far been found. Therefore, the RNA population inside a cell is diverse and cryptic and, thus, demands powerful methods to study its composition, abundance, and structure. DNA oligonucleotide microarrays have proven to be of great utility to study transcription of genes in various organisms. Recently, due to advancement in microarray technology, tiling microarrays that extend transcription measurement to genomic regions beyond protein-coding genes were designed for several species. The Saccharomyces cerevisiae yeast tiling array contains overlapping probes across the full genomic sequence, with consecutive probes starting every 8 bp on average on each strand, enabling strand-specific measurement of transcription from a full eukaryotic genome. Here, we describe the methods used to extract yeast RNA, convert it into first-strand cDNA, fragment, and label it for hybridization to the tiling array. This protocol will enable researchers not only to study which genes are expressed and to what levels, but also to identify non-coding RNAs and to study the structure of transcripts including their untranslated regions, alternative start, stop, and processing sites. This information will allow understanding their roles inside cells.

Original languageAmerican English
Title of host publicationYeast Systems Biology
Subtitle of host publicationMethods and Protocols
PublisherHumana Press Inc.
Pages107-123
Number of pages17
ISBN (Print)9781617791727
DOIs
StatePublished - 2011

Publication series

NameMethods in Molecular Biology
Volume759
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029

Bibliographical note

Publisher Copyright:
© 2011, Humana Press.

Keywords

  • Tiling microarray
  • cDNA
  • gene expression
  • gene structure
  • non-coding RNA
  • strand-specific transcription
  • transcription
  • whole-genome microarray
  • yeast

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