Nature Biotechnology | Research | Article
Published online13 October 2013 A single-molecule long-read survey of the human transcriptome
Abstract•Author information•Supplementary information Global RNA studies have become central to understanding biological processes, but methods such as microarrays and short-read sequencing are unable to describe an entire RNA molecule from 5' to 3' end. Here we use single-molecule long-read sequencing technology from Pacific Biosciences to sequence the polyadenylated RNA complement of a pooled set of 20 human organs and tissues without the need for fragmentation or amplification. We show that full-length RNA molecules of up to 1.5 kb can readily be monitored with little sequence loss at the 5' ends. For longer RNA molecules more 5' nucleotides are missing, but complete intron structures are often preserved. In total, we identify ~14,000 spliced GENCODE genes. High-confidence mappings are consistent with GENCODE annotations, but 10% of the alignments represent intron structures that were not previously annotated. As a group, transcripts mapping to unannotated regions have features of long, noncoding RNAs. Our results show the feasibility of deep sequencing full-length RNA from complex eukaryotic transcriptomes on a single-molecule level.
(WvSchaik) "Is there even a need for Nanopore sequencing now that PacBio has improved its technology so much in the last year or so?" Tweet 12:46 PM - 13 Oct 13 ·