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Nat Methods. 2014 Sep;11(9):959-65. doi: 10.1038/nmeth.3029. Epub 2014 Jul 13.

RNA motif discovery by SHAPE and mutational profiling (SHAPE-MaP).

Author information

  • 11] Department of Chemistry, University of North Carolina, Chapel Hill, North Carolina, USA. [2].
  • 2Department of Microbiology and Immunology, University of North Carolina, Chapel Hill, North Carolina, USA.
  • 3Department of Chemistry, University of North Carolina, Chapel Hill, North Carolina, USA.

Abstract

Many biological processes are RNA-mediated, but higher-order structures for most RNAs are unknown, which makes it difficult to understand how RNA structure governs function. Here we describe selective 2'-hydroxyl acylation analyzed by primer extension and mutational profiling (SHAPE-MaP) that makes possible de novo and large-scale identification of RNA functional motifs. Sites of 2'-hydroxyl acylation by SHAPE are encoded as noncomplementary nucleotides during cDNA synthesis, as measured by massively parallel sequencing. SHAPE-MaP-guided modeling identified greater than 90% of accepted base pairs in complex RNAs of known structure, and we used it to define a new model for the HIV-1 RNA genome. The HIV-1 model contains all known structured motifs and previously unknown elements, including experimentally validated pseudoknots. SHAPE-MaP yields accurate and high-resolution secondary-structure models, enables analysis of low-abundance RNAs, disentangles sequence polymorphisms in single experiments and will ultimately democratize RNA-structure analysis.

PMID:
25028896
PMCID:
PMC4259394
DOI:
10.1038/nmeth.3029
[PubMed - indexed for MEDLINE]
Free PMC Article
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