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Nat Methods DOI:10.1038/s41592-018-0014-2

Comprehensive comparative analysis of 5'-end RNA-sequencing methods.

Publication TypeJournal Article
Year of Publication2018
AuthorsAdiconis, X, Haber, AL, Simmons, SK, Moonshine, ALevy, Ji, Z, Busby, MA, Shi, X, Jacques, J, Lancaster, MA, Pan, JQ, Regev, A, Levin, JZ
JournalNat Methods
Date Published2018 07
KeywordsBase Sequence, Brain, Embryonic Stem Cells, Gene Library, Humans, RNA, Sequence Analysis, RNA

Specialized RNA-seq methods are required to identify the 5' ends of transcripts, which are critical for studies of gene regulation, but these methods have not been systematically benchmarked. We directly compared six such methods, including the performance of five methods on a single human cellular RNA sample and a new spike-in RNA assay that helps circumvent challenges resulting from uncertainties in annotation and RNA processing. We found that the 'cap analysis of gene expression' (CAGE) method performed best for mRNA and that most of its unannotated peaks were supported by evidence from other genomic methods. We applied CAGE to eight brain-related samples and determined sample-specific transcription start site (TSS) usage, as well as a transcriptome-wide shift in TSS usage between fetal and adult brain.


Alternate JournalNat Methods
PubMed ID29867192
PubMed Central IDPMC6075671
Grant ListU24 CA180922 / CA / NCI NIH HHS / United States
MC_UP_1201/9 / MRC_ / Medical Research Council / United Kingdom
R00 CA207865 / CA / NCI NIH HHS / United States
U01 MH105960 / MH / NIMH NIH HHS / United States
K99 CA207865 / CA / NCI NIH HHS / United States
R01 MH115045 / MH / NIMH NIH HHS / United States
RM1 HG006193 / HG / NHGRI NIH HHS / United States