High-Resolution Analysis of the 5′-End Transcriptome Using a Next Generation DNA Sequencer

• DOI: 10.1371/journal.pone.0004108
• Publication Date: 2008-12-31T23:07:23Z
• Authors: Shin-ichi Hashimoto; Wei Qu; Budrul Ahsan; Katsumi Ogoshi; Atsushi Sasaki; Yoichiro Nakatani; Yongjun Lee; Masako Ogawa; Akio Ametani; Yutaka Suzuki; Sumio Sugano; Clarence C. Lee; Robert C. Nutter; Shinichi Morishita; Kouji Matsushima
• Keywords: RefSeq; Sanger sequencing

Massively parallel, tag-based sequencing systems, such as the SOLiD system, hold promise of revolutionizing study whole genome gene expression due to number data points that can be generated in a simple and cost-effective manner. We describe development 5'-end transcriptome workflow for system demonstrate advantages sensitivity dynamic range offered by this application over traditional approaches expression. analysis was used within colon cancer cell line, HT-29, treated with DNA methyltransferase inhibitor, 5-aza-2'-deoxycytidine (5Aza). More than 20 million 25-base tags were obtained from untreated 5Aza-treated cells matched sequences human genome. Seventy three percent mapped unique associated RefSeq cDNA sequences, corresponding approximately 14,000 different protein-coding genes single type. The level these ranged 0.02 4,704 transcripts per cell. sequence run platform 100-1,000 fold greater observed 5'end SAGE 70,000 Sanger sequencing. high-resolution profiling presented will not only provide novel insight into transcriptional machinery but should also serve basis better understanding biology.