RNA-Protein Interaction Protocols

Due to the vital biological importance of RNA and proteins functioning together within a cell, a protocol volume describing experimental procedures to study their interactions should find a home in many laboratories. RNA-Protein Interaction Protocols, Second Edition updates, complements, and expands upon the popular first edition by providing a collection of cutting-edge techniques developed or refined in the past few years along with tried-and-true methods. The expert contributors explore the isolation and characterization of RNA-protein complexes, the analysis and measurement of RNA-protein interaction, and related novel techniques and strategies. Written in the highly successful Methods in Molecular Biology™ series format, the chapters include brief introductions to the material, lists of necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and a Notes section which highlights tips on troubleshooting and avoiding known pitfalls.

Comprehensive and up-to-date, RNA-Protein Interaction Protocols, Second Edition is an ideal guide for researchers continuing the study of this all-important biological partnership.

Various methods for affinity-purifying cellular or in vitro reconstituted RNPs A collection of methods for studying RNA-protein interaction occurred inside a cell Various methods for assaying protein footprints on RNA Various methods for mapping protein regions involved in RNA interaction New methods including quantifying specific RNAs, designing siRNAs, measuring RNPase activity, etc. Example methods in many cellular events

Inhalt
Isolation of a Sequence-Specific RNA Binding Protein, Polypyrimidine Tract Binding Protein, Using RNA Affinity Chromatography.- An Affinity Oligonucleotide Displacement Strategy to Purify Ribonucleoprotein Complexes Applied to Human Telomerase.- RNA Affinity Tags for the Rapid Purification and Investigation of RNAs and RNAProtein Complexes.- Assembly and Glycerol Gradient Isolation of Yeast Spliceosomes Containing Transcribed or Synthetic U6 snRNA.- Purification of Ribonucleoproteins Using Peptide-Elutable Antibodies and Other Affinity Techniques.- CLIP: Crosslinking and ImmunoPrecipitation of In Vivo RNA Targets of RNA-Binding Proteins.- Quantitative Analysis of Protein-RNA Interactions by Gel Mobility Shift.- Monitoring Assembly of Ribonucleoprotein Complexes by Isothermal Titration Calorimetry.- Characterization of RNAProtein Interactions by Phosphorothioate Footprinting and Its Applications to the Ribosome.- In Vivo Analysis of Ribonucleoprotein Complexes Using Nucleotide Analog Interference Mapping.- T7 RNA Polymerase-Mediated Incorporation of 8-N3AMP Into RNA for Studying Protein-RNA Interactions.- A Simple Crosslinking Method, CLAMP, to Map the Sites of RNA-Contacting Domains Within a Protein.- Proteins Specifically Modified With a Chemical Nuclease as Probes of RNA-Protein Interaction.- RNA-Protein Crosslink Mapping Using TEV Protease.- Structural Analysis of Protein-RNA Interactions With Mass Spectrometry.- Analyzing RNA-Protein Crosslinking Sites in Unlabeled Ribonucleoprotein Complexes by Mass Spectrometry.- In Vitro Selection of Random RNA Fragments to Identify Protein-Binding Sites Within Large RNAs.- Immunoprecipitation Analysis to Study RNA-Protein Interactions in Xenopus Oocytes.- Mapping the Regions of RNase P Catalytic RNA That Are Potentially inClose Contact With Its Protein Cofactor.- Quantification of MicroRNAs, Splicing Isoforms, and Homologous mRNAs With the Invader Assay.- Analysis of RNA Structure and RNA-Protein Interactions in Mammalian Cells by Use of Terminal Transferase-Dependent PCR.- Duplex Unwinding and RNP Remodeling With RNA Helicases.- Preparation of Efficient Splicing Extracts From Whole Cells, Nuclei, and Cytoplasmic Fractions.- Designing and Utilization of siRNAs Targeting RNA Binding Proteins.- The use of Saccharomyces cerevisiae proteomic libraries to identify RNA-modifying proteins.