Construction of Subtracted EST and Normalised cDNA Libraries From Liver of Chemical-Exposed Three-Spined Stickleback (Gasterosteus Aculeatus) Containing Pollutant-Responsive Genes as a Resource for Transcriptome Analysis

Mar Environ Res. 2008 Jul;66(1):127-30. doi: 10.1016/j.marenvres.2008.02.043. Epub 2008 Feb 26.

Abstract

The three-spined stickleback (Gasterosteus aculeatus) is ideally suited to laboratory studies, while its wide distribution in the northern hemisphere gives it great potential as a sentinel organism. In the setting of a UK-wide collaboration (Fish Toxicogenomics) we have developed a microarray for transcriptomic analysis of chemical responses in populations of G. aculeatus under laboratory and field conditions. Although several EST libraries are available for this species none are from chemical-exposed fish and thus unlikely to include a full set of pollutant-responsive genes. To harvest such transcripts cDNA libraries were produced from liver of chemical-exposed mature males. Two normalised full-length libraries were generated by different methods: (1) partial subtraction of polyA+ RNA against solid-phase cDNA using magnetic bead technology; (2) degradation of double stranded cDNA formed by abundant transcripts. To enrich for pollutant-responsive genes a subtracted EST library was also generated. For each library approximately 1.5K clones were sequenced and characterised using Blast2GO. All libraries contained pollutant-responsive transcripts not previously available while additionally the subtracted library was generally enriched approximately 1.2-10-fold for transcripts expected to be induced in response to the pollutants.

MeSH terms

  • Animals
  • Expressed Sequence Tags*
  • Gene Expression Profiling
  • Gene Expression Regulation / drug effects*
  • Gene Library*
  • Liver / drug effects*
  • Liver / metabolism*
  • Male
  • Molecular Sequence Data
  • Smegmamorpha / genetics*
  • Water Pollutants, Chemical / toxicity*

Substances

  • Water Pollutants, Chemical

Associated data

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