Identifying Novel Genes in C. elegans Using SAGE Tags

AbstractBackground: Despite extensive efforts devoted to predicting protein-coding genes in genome sequences, manybona fide genes have not been found and many existing gene models are not accurate in all sequenced eukaryotegenomes. This situation is partly explained by the fact that gene prediction programs have been developed basedon our incomplete understanding of gene feature information such as splicing and promoter characteristics.Additionally, full-length cDNAs of many genes and their isoforms are hard to obtain due to their low level or rareexpression. In order to obtain full-length sequences of all protein-coding genes, alternative approaches arerequired.Results: In this project, we have developed a method of reconstructing full-length cDNA sequences based onshort expressed sequence tags which is called sequence tag-based amplification of cDNA ends (STACE). Expressedtags are used as anchors for retrieving full-length transcripts in two rounds of PCR amplification. We havedemonstrated the application of STACE in reconstructing full-length cDNA sequences using expressed tags minedin an array of serial analysis of gene expression (SAGE) of C. elegans cDNA libraries. We have successfully appliedSTACE to recover sequence information for 12 genes, for two of which we found isoforms. STACE was used tosuccessfully recover full-length cDNA sequences for seven of these genes.Conclusions: The STACE method can be used to effectively reconstruct full-length cDNA sequences of genes thatare under-represented in cDNA sequencing projects and have been missed by existing gene prediction methods,but their existence has been suggested by short sequence tags such as SAGE tags.