Expressed sequence tags: normalization and subtraction of cDNA libraries expressed sequence tags\ normalization and subtraction of cDNA libraries.

Marcelo Bento Soares, Maria de Fatima Bonaldo, Jeremiah D. Hackett, Debashish Bhattacharya

Research output: Contribution to journalArticle

Abstract

Expressed Sequence Tags (ESTs) provide a rapid and efficient approach for gene discovery and analysis of gene expression in eukaryotes. ESTs have also become particularly important with recent expanded efforts in complete genome sequencing of understudied, nonmodel eukaryotes such as protists and algae. For these projects, ESTs provide an invaluable source of data for gene identification and prediction of exon-intron boundaries. The generation of EST data, although straightforward in concept, requires nonetheless great care to ensure the highest efficiency and return for the investment in time and funds. To this end, key steps in the process include generation of a normalized cDNA library to facilitate a high gene discovery rate followed by serial subtraction of normalized libraries to maintain the discovery rate. Here we describe in detail, protocols for normalization and subtraction of cDNA libraries followed by an example using the toxic dinoflagellate Alexandrium tamarense.

Original languageEnglish (US)
Pages (from-to)109-122
Number of pages14
JournalMethods in molecular biology (Clifton, N.J.)
Volume533
StatePublished - 2009
Externally publishedYes

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

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