Goldberg Martin W. (ed.) The Nuclear Pore: Complex Methods and Protocols

Humana Press, 2022. — 506 p. — (Methods in Molecular Biology 2502). — ISBN 978-1-0716-2336-7.This volume covers all methods used to discover the composition, structure, and dynamics of the nuclear pore complex (NPC), as well as the soluble transport factors involved in the transport process. Chapters detail structural biology methods, biophysical, transport factors , and a wide range of model organisms that have been used over the years. Written in the format of the highly successful Methods in Molecular Biology series, each chapter includes an introduction to the topic, lists necessary materials and reagents, includes tips on troubleshooting and known pitfalls, and step-by-step, readily reproducible protocols. Authoritative and cutting-edge, The Nuclear Pore Complex: Methods and Protocols: Methods and Protocols aims to be a useful and practical guide to new researchers and experts looking to expand their knowledge.The Nuclear Pore Complex and Nucleoporins
Affinity Isolation of Endogenous Saccharomyces Cerevisiae Nuclear Pore Complexes
Transformation of Chaetomium thermophilum and Affinity Purification of Native Thermostable Protein Complexes
Nuclear Pore Complex Assembly Using Xenopus Egg Extract
Nucleo-cytoplasmic Passage
Analysis of Nuclear Pore Complex Permeability in Mammalian Cells and Isolated Nuclei Using Fluorescent Dextrans
Hormone-Inducible Transport Reporter Assay to Study Nuclear Import Defects in Neurodegenerative Diseases
Subcellular Fractionation Suitable for Studies of RNA and Protein Trafficking
Localizing Total mRNA in Plant Cells
Using Single Molecule RNA FISH to Determine Nuclear Export and Transcription Phenotypes in Drosophila Tissues
Functional Analysis of Nucleoporins
Analysis of Nucleoporin Function Using Inducible Degron Techniques
Monitoring of Chromatin Organization at the Nuclear Pore Complex, Inner Nuclear Membrane, and Nuclear Interior in Live Cells by Fluorescence Ratiometric Imaging of Chromatin (FRIC).
Analysis of Nuclear Pore Complexes in Caenorhabditis elegans by Live Imaging and Functional Genomics
Protein Retargeting in Aspergillus nidulans to Study the Function of Nuclear Pore Complex Proteins
Protein–Protein Interactions
Split-GFP Complementation to Study the Nuclear Membrane Proteome Using Microscopy
Bimolecular Fluorescence Complementation: Quantitative Analysis of In Cell Interaction of Nuclear Transporter Importin α with Cargo Proteins
Validation of Nuclear Pore Complex Protein–Protein Interactions by Transient Expression in Plants
Protein–Protein Interactions
Binding Affinity Measurement of Nuclear Export Signal Peptides to Their Exporter CRM1
Post Translational Modifications
Analysis of Ubiquitylation and SUMOylation of Yeast Nuclear Pore Complex Proteins
Purification of Cdk-CyclinB-Kinase–Targeted Phosphopeptides from Nuclear Envelope
Biophysical Methods
Crystallization of Nuclear Export Signals or Small-Molecule Inhibitors Bound to Nuclear Exporter CRM1
Atomic Force Microscopy for Structural and Biophysical Investigations on Nuclear Pore Complexes
Multivalent Interactions with Intrinsically Disordered Proteins Probed by Surface Plasmon Resonance
Assembly and Use of a Microfluidic Device to Study Nuclear Mechanobiology During Confined Migration
Imaging NPCs and Transport
Speed Microscopy: High-Speed Single Molecule Tracking and Mapping of Nucleocytoplasmic Transport
Imaging Fluorescent Nuclear Pore Complex Proteins in C. elegans
Visualizing Nuclear Pore Complexes in Xenopus Egg Extracts
TEM Imaging of Membrane Choreography During Mitosis of Drosophila Tissue Culture Cells
Scanning Electron Microscopy (SEM) and Immuno-SEM of Nuclear Pore Complexes from Amphibian Oocytes, Mammalian Cell Cultures, Yeast, and Plants
NPC Structure in Model Organisms: Transmission Electron Microscopy and Immunogold Labeling Using High-Pressure Freezing/Freeze Substitution of Yeast, Worms, and Plants
High-Resolution Imaging and Analysis of Individual Nuclear Pore Complexes
Live CLEM Imaging of Tetrahymena to Analyze the Dynamic Behavior of the Nuclear Pore Complex
Visualizing Nuclear Pore Complex Assembly In Situ in Human Cells at Nanometer Resolution by Correlating Live Imaging with Electron Microscopy