The second edition of a highly acclaimed handbook and ready reference. Unmatched in its breadth and quality, around 100 specialists from all over the world share their up-to-date expertise and experiences, including hundreds of protocols, complete with explanations, and hitherto unpublished troubleshooting hints. They cover all modern techniques for the handling, analysis and modification of RNAs and their complexes with proteins. Throughout, they bear the practising bench scientist in mind, providing quick and reliable access to a plethora of solutions for practical questions of RNA research, ranging from simple to highly complex. This broad scope allows the treatment of specialized methods side by side with basic biochemical techniques, making the book a real treasure trove for every researcher experimenting with RNA.

Table of Contents:
Preface XXXV List of Contributors XXXVII Part I RNA Synthesis and Detection 1 1 Enzymatic RNA Synthesis Using Bacteriophage T7 RNA Polymerase 3 Markus Gößringer, Dominik Helmecke, Karen Köhler, Astrid Schön, Leif A. Kirsebom, Albrecht Bindereif, and Roland K. Hartmann 2 Production of RNAs with Homogeneous 5'- and 3'-Ends 29 Mario Mörl and Roland K. Hartmann 3 RNA Ligation 45 Janne J. Turunen, Liudmila V. Pavlova, Martin Hengesbach, Mark Helm, Sabine Müller, Roland K. Hartmann, and Mikko J. Frilander 4 Northern Blot Detection of Small RNAs 89 Benedikt M. Beckmann, Arnold Grünweller, and Roland K. Hartmann 5 Rapid, Non-Denaturing, Large-Scale Purification of In Vitro Transcribed RNA Using Weak Anion-Exchange Chromatography 105 Laura E. Easton, Yoko Shibata, and Peter J. Lukavsky 6 3'-Terminal Attachment of Fluorescent Dyes and Biotin 117 Dagmar K. Willkomm and Roland K. Hartmann 7 Chemical RNA Synthesis, Purification, and Analysis 129 Brian S. Sproat 8 Modified RNAs as Tools in RNA Biochemistry 151 Thomas E. Edwards and Snorri Th. Sigurdsson Part II Structure Determination 173 9 Direct Determination of RNA Sequence and Modification by Radiolabeling Methods 175 Olaf Gimple and Astrid Schön 10 Probing RNA Structure In Vitro with Enzymes and Chemicals 205 Anne-Catherine Helfer, Cédric Romilly, Clément Chevalier, Efthimia Lioliou, Stefano Marzi, and Pascale Romby 11 Probing RNA Solution Structure by Photocrosslinking: Incorporation of Photoreactive Groups at RNA Termini and Determination of Crosslinked Sites by Primer Extension 231 Michael E. Harris 12 Terbium(III) Footprinting as a Probe of RNA Structure and Metal Binding Sites 255 Dinari A. Harris, Gabrielle C. Todd, and Nils G. Walter 13 Pb2+-Induced Cleavage of RNA 269 Leif A. Kirsebom and Jerzy Ciesiolka 14 Identification and Characterization of Metal Ion Coordination Interactions with RNA by Quantitative Analysis of Thiophilic Metal Ion Rescue of Site-Specific Phosphorothioate Modifications 285 Michael E. Harris 15 Probing RNA Structure and Ligand Binding Sites on RNA by Fenton Cleavage 301 Corina G. Heidrich and Christian Berens 16 Measuring the Stoichiometry of Magnesium Ions Bound to RNA 319 Andrew J. Andrews and Carol A. Fierke 17 Nucleotide Analog Interference Mapping and Suppression (NAIM/NAIS): a Combinatorial Approach to Study RNA Structure, Folding, and Interaction with Proteins 329 Olga Fedorova, Marc Boudvillain, and Christina Waldsich 18 Nucleotide Analog Interference Mapping (NAIM): Application to the RNase P System 369 Simona Cuzic-Feltens and Roland K. Hartmann 19 Identification of Divalent Metal Ion Binding Sites in RNA/DNA-Metabolizing Enzymes by Fe(II)-Mediated Hydroxyl Radical Cleavage 397 Yan-Guo Ren, Niklas Henriksson, and Anders Virtanen 20 RNA Structure and Folding Analyzed Using Small-Angle X-Ray Scattering 407 Nathan J. Baird, Jeremey West, and Tobin R. Sosnick 21 Temperature-Gradient Gel Electrophoresis of RNA 427 Detlev Riesner and Gerhard Steger 22 UV Melting Studies with RNA 445 Philippe Dumas, Eric Ennifar, Francois Disdier, and Philippe Walter 23 RNA Crystallization 481 Jiro Kondo, Claude Sauter, and Benôýt Masquida 24 Studying RNA Using Single Molecule Fluorescence Resonance Energy Transfer 499 Felix Spenkuch, Olwen Domingo, Gerald Hinze, Thomas Basché, and Mark Helm 25 Atomic Force Microscopy Imaging and Force Spectroscopy of RNA 527 Malte Bussiek, Antonie Schöne, and Wolfgang Nellen Part III RNA Genomics & Bioinformatics, Global Approaches 547 26 Secondary Structure Prediction 549 Gerhard Steger 27 RNA Secondary Structure Analysis Using Abstract Shapes 579 Robert Giegerich and Björn Voß 28 Screening Genome Sequences for known RNA Genes or Motifs 595 Daniel Gautheret 29 Homology Search for Small Structured Non-coding RNAs 619 Manja Marz, Stefanie Wehner, and Peter F. Stadler 30 Predict RNA 2D and 3D Structure over the Internet Using MC-Tools 633 Stephen Leong Koan, Jonathan Roy, Marc Parisien, and Francois Major 31 S2S-Assemble2: a Semi-Automatic Bioinformatics Framework to Study and Model RNA 3D Architectures 667 Fabrice Jossinet and Eric Westhof 32 Molecular Dynamics Simulations of RNA Systems 687 Pascal Auffinger 33 Identification and Characterization of Small Non-coding RNAs in Bacteria 719 Dimitri Podkaminski, Marie Bouvier, and Jörg Vogel 34 The Identification of Bacterial Non-coding RNAs through Complementary Approaches 787 Björn Voß and Wolfgang R. Hess 35 Experimental RNomics, a Global Approach to Identify Non-coding RNAs in Model Organisms, and RNPomics to Analyze the Non-coding RNP Transcriptome 801 Mathieu Rederstorff and Alexander Hüttenhofer 36 Computational Methods for Gene Expression Profiling Using Next-Generation Sequencing (RNA-Seq) 821 John C. Castle 37 Characterization and Prediction of miRNA Targets 833 Jean Hausser and Mihaela Zavolan 38 Barcoded cDNA Libraries for miRNA Profiling by Next-Generation Sequencing 861 Markus Hafner, Neil Renwick, John Pena, Aleksandra Mihailovic, and Thomas Tuschl 39 Transcriptome-Wide Identification of Protein Binding Sites on RNA by PAR-CLIP (Photoactivatable-Ribonucleoside-Enhanced Crosslinking and Immunoprecipitation) 877 Jessica I. Hoell, Markus Hafner, Markus Landthaler, Manuel Ascano, Thalia A. Farazi, Greg Wardle, Jeff Nusbaum, Pavol Cekan, Mohsen Khorshid, Lukas Burger, Mihaela Zavolan, and Thomas Tuschl 40 Global Analysis of Protein–RNA Interactions with Single-Nucleotide Resolution Using iCLIP 899 Julian König, Nicholas J. Mc Glincy, and Jernej Ule Part IV RNA Function, RNP Analysis, SELEX, RNAi 919 41 Use of RNA Affinity Matrices for the Isolation of RNA Binding Proteins 921 Markus Englert, Bettina Späth, Steffen Schiffer, Sylvia Rösch, Hildburg Beier, and Anita Marchfelder 42 Biotin-Based Affinity Purification of RNA–Protein Complexes 935 Marco Preußner, Silke Schreiner, Inna Grishina, Zsofia Palfi, Jingyi Hui, and Albrecht Bindereif 43 Affinity Purification of Spliceosomal and Small Nuclear Ribonucleoprotein Complexes 957 Julia Dannenberg, Patrizia Fabrizio, Cindy L. Will, and Reinhard Lührmann 44 Study of RNA–Protein Interactions and RNA Structure in Ribonucleoprotein Particles (RNPs) 975 Virginie Marchand, Annie Mougin, Agn´es Méreau, Isabelle Behm-Ansmant, Yuri Motorin, and Christiane Branlant 45 Immunopurification of Endogenous RNAs Associated with RNA Binding Proteins In vivo 1017 Minna-Liisa Änkö and Karla M. Neugebauer 46 Protein–RNA Crosslinking in Native Ribonucleoprotein Particles 1029 Olexandr Dybkov, Henning Urlaub, and Reinhard Lührmann 47 Sedimentation Analysis of Ribonucleoprotein Complexes 1055 Tanja Rösel, Jan Medenbach, Andrey Damianov, Silke Schreiner, and Albrecht Bindereif 48 Identification and Characterization of RNA Binding Proteins through Three-Hybrid Analysis 1067 Felicia Scott and David R. Engelke 49 Experimental Identification of MicroRNA Targets 1087 Michaela Beitzinger and Gunter Meister 50 Aptamer Selection against Biological Macromolecules: Proteins and Carbohydrates 1097 Franziska Peter and C. Stefan Voertler 51 In Vitro Selection against Small Targets 1139 Dirk Eulberg, Christian Maasch, Werner G. Purschke, and Sven Klussmann 52 SELEX Strategies to Identify Antisense and Protein Target Sites in RNA or hnRNP Complexes 1165 Martin Lützelberger, Martin R. Jakobsen, and Jørgen Kjems 53 Genomic SELEX 1185 Jennifer L. Boots, Katarzyna Matylla-Kulinska, Marek Zywicki, Bob Zimmermann, and Renée Schroeder 54 In vivo SELEX Strategies 1207 Thomas A. Cooper 55 Gene Silencing Methods Using Vector-Encoded siRNAs or miRNAs 1221 Ying Poi Liu and Ben Berkhout 56 Using Chemical Modification to Enhance siRNA Performance 1243 Jesper B. Bramsen, Arnold Grünweller, Roland K. Hartmann, and Jørgen Kjems Appendix: UV Spectroscopy for the Quantitation of RNA 1279 Index 1283