Tag-based amplification of cDNA ends: a new approach to gene discovery

Gene prediction programs, though effective, produce both false positive and false negative predictions. Transcriptional evidence from cDNA and EST sequences will not produce false positive gene models and provide the structure of a gene. However, this approach is hampered by high cost and the inability to sequence genes that have low expression. New methods that are low in cost and are capable of providing sequence for transiently expressed genes are needed. This thesis describes a method that targets undiscovered polyadenylated transcripts for characterization. Tag-based amplification of cDNA ends (TACE) consists of two components. First, sequence tags (i.e. SAGE tags) that could represent novel genes are selected. Second, the sequence found upstream and downstream of the tag is amplified via PCR. Resulting sequences are aligned for gene model characterization. TACE has produced evidence for six new genes, four annotation extensions, and two new gene models for a pseudogene and ncRNA.