Traditional Chemical Mapping of RNA Structure In Vitro and In Vivo.
Chemical probing is often used to gain knowledge on the secondary and tertiary structures of RNA molecules either free or engaged in complexes with ligands. The method monitors the reactivity of each nucleotide towards chemicals of various specificities reflecting the hydrogen bonding environment of each nucleotide within the RNA molecule. In addition, information can be obtained on the binding site of a ligand (noncoding RNAs, protein, metabolites), and on RNA conformational changes that accompanied ligand binding or perturbation of the environmental cues. The detection of the modifications can be obtained either by using end-labeled RNA molecules or by primer extension using reverse transcriptase. The goal of this chapter is to provide the reader with an experimental guide to probe the structure of RNA in vitro and in vivo with the most suitable chemical probes.
Biotechnologie et Signalisation Cellulaire, CNRS-INSERM, ESBS, Université de Strasbourg, 300 boulevard Sebastien Brant, Illkirch, 67412, France.
Pierre
P
Fechter
Architecture et Réactivité de l'ARN, CNRS, IBMC, Université de Strasbourg, 15 rue René Descartes, 67084, Strasbourg, France.
Delphine
D
Parmentier
College of Veterinary Medicine, Nanjing Agricultural University, 1 Weigang Road, Nanjing, 210095, China.
ZongFu
Z
Wu
Architecture et Réactivité de l'ARN, CNRS, IBMC, Université de Strasbourg, 15 rue René Descartes, 67084, Strasbourg, France.
Olivier
O
Fuchsbauer
Architecture et Réactivité de l'ARN, CNRS, IBMC, Université de Strasbourg, 15 rue René Descartes, 67084, Strasbourg, France. P.romby@ibmc-cnrs.unistra.fr.
Pascale
P
Romby
Architecture et Réactivité de l'ARN, CNRS, IBMC, Université de Strasbourg, 15 rue René Descartes, 67084, Strasbourg, France.
Stefano
S
Marzi
D019295:Computational Biology / Q000379:methods*; D004591:Electrophoresis, Polyacrylamide Gel; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D012313:RNA / Q000737:chemistry*; D059367:RNA Cleavage; D059370:RNA Folding
D012313:RNA
Chemical mapping; Lead(II) induced cleavages; RNA structure probing