Research article

Transcriptome analysis for Caenorhabditis elegans based on novel expressed sequence tags

Heesun Shin^1,2 , Martin Hirst² , Matthew N Bainbridge² , Vincent Magrini³ , Elaine Mardis³ , Donald G Moerman⁴ , Marco A Marra² , David L Baillie¹ and Steven JM Jones²

¹Molecular Biology and Biochemistry, Simon Fraser University, Burnaby, BC, Canada

²Canada's Michael Smith Genome Sciences Centre, British Columbia Cancer Research Centre, British Columbia Cancer Agency, Vancouver, BC, Canada

³Genome Sequencing Center, Washington University School of Medicine, St Louis, USA

⁴Department of Zoology, University of British Columbia, Vancouver, BC, Canada

author email corresponding author email

BMC Biology 2008, 6:30doi:10.1186/1741-7007-6-30

Published:	8 July 2008

Abstract

Background

We have applied a high-throughput pyrosequencing technology for transcriptome profiling of Caenorhabditis elegans in its first larval stage. Using this approach, we have generated a large amount of data for expressed sequence tags, which provides an opportunity for the discovery of putative novel transcripts and alternative splice variants that could be developmentally specific to the first larval stage. This work also demonstrates the successful and efficient application of a next generation sequencing methodology.

Results

We have generated over 30 million bases of novel expressed sequence tags from first larval stage worms utilizing high-throughput sequencing technology. We have shown that approximately 14% of the newly sequenced expressed sequence tags map completely or partially to genomic regions where there are no annotated genes or splice variants and therefore, imply that these are novel genetic structures. Expressed sequence tags, which map to intergenic (around 1000) and intronic regions (around 580), may represent novel transcribed regions, such as unannotated or unrecognized small protein-coding or non-protein-coding genes or splice variants. Expressed sequence tags, which map across intron-exon boundaries (around 300), indicate possible alternative splice sites, while expressed sequence tags, which map near the ends of known transcripts (around 600), suggest extension of the coding or untranslated regions. We have also discovered that intergenic and intronic expressed sequence tags, which are well conserved across different nematode species, are likely to represent non-coding RNAs. Lastly, we have incorporated available serial analysis of gene expression data generated from first larval stage worms, in order to predict novel transcripts that might be specifically or predominantly expressed in the first larval stage.

Conclusion

We have demonstrated the use of a high-throughput sequencing methodology to efficiently produce a snap-shot of transcriptional activities occurring in the first larval stage of C. elegans development. Such application of this new sequencing technique allows for high-throughput, genome-wide experimental verification of known and novel transcripts. This study provides a more complete C. elegans transcriptome profile and, furthermore, gives insight into the evolutionary and biological complexity of this organism.