Structures of Large RNA Molecules and Their Complexes / edited by Sarah A. Woodson, Department of Biophysics, Johns Hopkins University, USA, Frédéric H.T. Allain, Institute of Molecular Biology and Biophysics, ETH Zürich, Switzerland.
Contributor(s): Woodson, Sarah A [editor.] | Allain, Frédéric H. T [editor.]Material type: TextSeries: Methods in enzymology: v. 558.Publisher: Waltham, MA : Academic Press, 2015Edition: First editionDescription: 1 online resource (xxi, 631 pages, 20 unnumbered pages of plates : illustrations (some color))Content type: text Media type: computer Carrier type: online resourceISBN: 9780128019368; 0128019360Subject(s): RNA | SCIENCE -- Life Sciences -- Biochemistry | RNA | RNA -- ultrastructureGenre/Form: Electronic books. Additional physical formats: Print version:: Structures of Large RNA Molecules and Their Complexes.DDC classification: 572.8/8 LOC classification: QP623 | .S77 2015ebOnline resources: Click here to access online
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Text in English.
This new volume of Methods in Enzymology continues the legacy of this premier serial with quality chapters authored by leaders in the field. This volume covers research methods in RNA folding and dynamics, RNA-protein interactions and large RNPs.
880-01 Front Cover; Structures of Large RNA Molecules and Their Complexes; Copyright; Contents; Contributors; Preface; Section I: RNA Structure and Dynamics; Chapter 1: Native Purification and Analysis of Long RNAs; 1. Introduction; 2. Native Purification of Long Noncoding RNAs; 2.1. Construct design; 2.2. DNA plasmid linearization; 2.3. In vitro transcription of RNA; 2.4. DNase digestion; 2.5. Proteinase K treatment; 2.6. EDTA chelation of divalent ions (optional); 2.7. Buffer exchange and purification; 2.8. Size-exclusion chromatography.
3. Study of the RNA Tertiary Folding by Sedimentation Velocity Analytical Ultracentrifugation3.1. Preparation of samples for a study of RNA folding; 3.2. Assembly of the optical cells, sample loading, and instrument setup; 3.3. Setting up a sedimentation velocity experiment; 3.4. Data analysis; 4. Analysis of the RNA Tertiary Folding by Analytical Size-Exclusion Chromatography; 5. Determination of the Secondary Structure of LncRNAs by Chemical Probing; 5.1. Designing and coupling of primers; 5.2. Generation of sequencing ladders; 5.3. SHAPE reaction; 5.4. DMS reaction.
5.5. Primer extension reaction5.6. Reactions for mobility shift correction; 5.7. Spectral calibration of the instrument; 5.8. Preparation of samples for capillary electrophoresis; 5.9. Data analysis; 5.9.1. Determination of chemical probing reactivity profiles; 5.9.2. Normalization of SHAPE and DMS reactivity profiles; 5.9.3. RNA secondary structure prediction and analysis; Acknowledgments; References; Chapter 2: Characterizing RNA Excited States Using NMR Relaxation Dispersion; 1. Introduction; 2. NMR Relaxation Dispersion; 2.1. Chemical Exchange; 2.2. RD Experiments.
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