title: Comparison of SHAPE reagents for mapping RNA structures inside living cells.
abstract: Recent advances in SHAPE technology have converted the classic primer extension method to next-generation sequencing platforms, allowing transcriptome-level analysis of RNA secondary structure. In particular, icSHAPE and SHAPE-MaP, using NAI-N3and 1M7 reagents, respectively, are methods that claim to measure in vivo structure with high-throughput sequencing. However, these compounds have not been compared on an unbiased, raw-signal level. Here, we directly compare several in vivo SHAPE acylation reagents using the simple primer extension assay. We conclude that while multiple SHAPE technologies are effective at measuring purified RNAs in vitro, acylimidazole reagents NAI and NAI-N3give markedly greater signals with lower background than 1M7 for in vivo measurement of the RNA structurome.
authors.affiliation: Center for Personal Dynamic Regulomes, Stanford University, Stanford, California 94305, USA.
authors.forename: Byron
authors.initials: B
authors.lastname: Lee
authors.affiliation: Center for Personal Dynamic Regulomes, Stanford University, Stanford, California 94305, USA.
authors.forename: Ryan A
authors.identifier: 0000-0001-5013-0442
authors.initials: RA
authors.lastname: Flynn
authors.affiliation: Department of Chemistry, Stanford University, Stanford, California 94305, USA.
authors.forename: Anastasia
authors.initials: A
authors.lastname: Kadina
authors.affiliation: Center for Personal Dynamic Regulomes, Stanford University, Stanford, California 94305, USA.
authors.forename: Jimmy K
authors.initials: JK
authors.lastname: Guo
authors.affiliation: Department of Chemistry, Stanford University, Stanford, California 94305, USA.
authors.forename: Eric T
authors.initials: ET
authors.lastname: Kool
authors.affiliation: Center for Personal Dynamic Regulomes, Stanford University, Stanford, California 94305, USA.
authors.forename: Howard Y
authors.initials: HY
authors.lastname: Chang
mesh_terms: D000215:Acylation; D000818:Animals; D001681:Biological Assay / Q000592:standards*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D059014:High-Throughput Nucleotide Sequencing; D007093:Imidazoles / Q000737:chemistry*; D007202:Indicators and Reagents; D051379:Mice; D000066450:Mouse Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism; D009602:Nitrosamines / Q000737:chemistry; D009690:Nucleic Acid Conformation; D010078:Oxazines / Q000737:chemistry*; D061251:Primary Cell Culture; D012313:RNA / Q000737:chemistry*
chemical_list: D007093:Imidazoles; D007202:Indicators and Reagents; D009602:Nitrosamines; D010078:Oxazines; C049700:methylisoamylnitrosamine; D012313:RNA; C037902:isatoic anhydride
keywords: RNA structure; SHAPE