Mapping of transcription start sites in Saccharomyces cerevisiae using 5′ SAGE

Abstract

A minimally addressed area in Saccharomyces cerevisiae research is the mapping of transcription start sites (TSS). Mapping of TSS in S.cerevisiae has the potential to contribute to our understanding of gene regulation, transcription, mRNA stability and aspects of RNA biology. Here, we use 5′ SAGE to map 5′ TSS in S.cerevisiae. Tags identifying the first 15-17 bases of the transcripts are created, ligated to form ditags, amplified, concatemerized and ligated into a vector to create a library. Each clone sequenced from this library identifies 10-20 TSS. We have identified 13 746 unique, unambiguous sequence tags from 2231 S.cerevisiae genes. TSS identified in this study are consistent with published results, with primer extension results described here, and are consistent with expectations based on previous work on transcription initiation. We have aligned the sequence flanking 4637 TSS to identify the consensus sequence A(Arich) 5NPyA(A/T)NN(Arich)6, which confirms and expands the previous reported PyA(A/T)Pu consensus pattern. The TSS data allowed the identification of a previously unrecognized gene, uncovered errors in previous annotation, and identified potential regulatory RNAs and upstream open reading frames in 5′-untranslated region. © The Author 2005. Published by Oxford University Press. All rights reserved.