RNA Detection

This volume introduces different concepts and methods of detecting RNA in biological material in a variety of model systems. The chapters in this book discuss methods that will answer numerous biological questions that arise in the study of RNAs. Some of the topics covered in this book are single mRNA molecule detection in embryos and neurons ; detection of mRNA and associated molecules by ISH-IEM on frozen sections; optimizing molecular beacons for intracellular analysis of RNA; imaging translation dynamics of single mRNA molecules in live cells; preparation of high-throughput sequencing libraries; and capturing RNA binding proteins in embryos and in cell-culture. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.

Cutting-edge and comprehensive, RNA Detection: Methods and Protocols is a valuable resource for novel and experiences scientist in the expanding field of RNAs.

Includes cutting-edge methods and protocols Provides step-by-step detail essential for reproducible results Contains key notes and implementation advice from the experts

Inhalt

The Secret Life of RNA: Lessons from Emerging Methodologies.- Quantification of 2'-O-Me Residues in RNA using Next-Generation Sequencing (Illumina RiboMethSeq Protocol).- Identifying the m 6 A Methylome by Affinity Purification and Sequencing.- PARIS: Psoralen Analysis of RNA Interactions and Structures with High Throughput and Resolution.- Axon-TRAP-RiboTag: Affinity-Purification of Translated mRNAs from Neuronal Axons in Mouse In Vivo.- LCM-Seq: A Method for Spatial Transcriptomic Profiling using Laser Capture Microdissection Coupled with PolyA-Based RNA Sequencing.- Spatial Transcriptomics: Constructing a Single-Cell Resolution Transcriptome-Wide Expression Atlas.- Single mRNA Molecule Detection in Drosophila.- Detection and Automated Analysis of Single Transcripts at Subcellular Resolution in Zebrafish Embryos.- Super-Resolution Single Molecule FISH at the Drosophila Neuromuscular Junction.- Detection of mRNA and Associated Molecules by ISH-IEM on Frozen Sections.- Hybridization Chain Reaction for Direct mRNA Detection without Nucleic Acid Purification.- In Situ Detection of MicroRNA Expression with RNAscope Probes.- Padlock Probes to Detect Single Nucleotide Polymorphisms.- Quantifying Gene Expression in Living Cells with Ratiometric Bimolecular Beacons.- Optimizing Molecular Beacons for Intracellular Analysis of RNA.- Live Imaging of Nuclear RNPs in Mammalian Complex Tissue with ECHO-liveFISH.- In Vivo Visualization and Function Probing of Transport mRNPs using Injected FIT Probes.- Visualizing RNA in Live Bacterial Cells using Fluorophore- and Quencher-Binding Aptamers.- Method for Imaging Live-Cell RNA using an RNA Aptamer and a Fluorescent Probe.- RNA Live Imaging in the Model Microorganism Ustilago maydis.- Real-Time Fluorescence Imaging of Single-Molecule Endogenous Non-Coding RNA in Living Cells.- Live Imaging of mRNA Synthesis in Drosophila.- Imaging Newly Transcribed RNA in Cells by Using a Clickable Azide-Modified UTP Analog.- Detection of the First Round of Translation: The TRICK Assay.- Imaging Translation Dynamics of Single mRNA Molecules in Live Cells.- Systematic Detection of Poly(A) + RNA-Interacting Proteins and Their Differential Binding.- Isolation and Characterization of Endogenous RNPs from Brain Tissues.- Individual Nucleotide Resolution UV Cross-Linking and Immunoprecipitation (iCLIP) to Determine Protein-RNA Interactions.- RNA Tagging: Preparation of High-Throughput Sequencing Libraries.- RAP-MS: A Method to Identify Proteins that Interact Directly with a Specific RNA Molecule in Cells.