Includes cutting-edge methods and protocols
Provides step-by-step detail essential for reproducible results
Contains key notes and implementation advice from the experts
This volume focuses on cytological, biochemical, and molecular biological methods to identify and examine the function of each nuclear body, with an emphasis on the analysis of long non-coding RNAs. Chapters focus on exploring recent studies that reveal how certain long non protein-coding RNAs accumulate in specific nuclear bodies and regulate the function of the bodies by serving as architectural components or controlling the dynamics of associating protein components. Written in the highly successful Methods of 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 key tips on troubleshooting and avoiding known pitfalls.
Authoritative and practical, Nuclear Bodies and Noncoding RNAs: Methods and Protocols serves as a guideline for further study into the sophisticated regulation of gene expression.
Part I: Imaging of Noncoding RNAs and Nuclear Bodies
1. Visualization of lncRNA by Single Molecule Fluorescence in situ Hybridization
Margaret Dunagin, Moran N. Cabili, John Rinn, and Arjun Raj
2. Super-resolution Imaging of Nuclear Bodies by STED Microscopy
Yasushi Okada and Shinichi Nakagawa
3. High-resolution 3D DNA FISH Using Plasmid Probes and Computational Correction of Optical Aberrations to Study Chromatin Structure at the Sub-megabase Scale
Luca Giorgetti, Tristan Piolot, and Edith Heard
4. Time-lapse Imaging of Nuclear Bodies
Saskia Hutten, Samuel Swift, and Angus I. Lamond
5. Visualization of Nucleic Acids with Synthetic Exciton-controlled Fluorescent Oligonucleotide Probes
Dan Ohtan Wang and Akimitsu Okamoto
6. Live CLEM Imaging to Analyze Nuclear Structures at High-resolution
Tokuko Haraguchi, Hiroko Osakada, and Takako Koujin
7. Ultrastructural Analysis of Nuclear Bodies Using Electron Microscopy
Sylvie Souquere and Gérard Pierron
8. Analyses of Nuclear Proteins and Nucleic Acid Structures Using Atomic Force Microscopy
Jamie L. Gilmore, Aiko Yoshida, Hirohide Takahashi, Katashi Deguchi, Toshiro Kobori, Emilie Louvet, Masahiro Kumeta, Shige H. Yoshimura, and Kunio Takeyasu
9. Genome-wide Co-localization Screening of Nuclear Body Components Using a Fluorescently Tagged FLJ cDNA Clone Library
Tetsuro Hirose and Naoki Goshima
Part II: Biochemical Analysis of Noncoding RNAs and Nuclear Bodies
10. Purification of Specific Chromatin Regions Using Oligonucleotides: Capture Hybridization Analysis of RNA Targets (CHART)
Christopher P. Davis and Jason A. West
11. RNA Antisense Purification (RAP) for Mapping RNA Interactions with Chromatin
Jesse Engreitz, Eric S. Lander, and Mitchell Guttman
12. In situ Dissection of RNA Functional Subunits by Domain-specific Chromatin Isolation by RNA Purification (dChIRP)
Jeffrey J. Quinn and Howard Y. Chang
13. Extracting, Enriching and Identifying Nuclear Body Sub-complexes Using Label-based Quantitative Mass Spectrometry
Archa Fox, Virja Mehta, Severine Boulon, and Laura Trinkle-Mulcahy
14. Studying RNA-binding Protein Interactions with Target mRNAs in Eukaryotic Cells: Native Ribonucleoprotein Immunoprecipitation (RIP) Assays
Joseph A. Cozzitorto, Maya Jimbo, Saswati Chand, Fernando Blanco, Shruti Lal, Melissa Gilbert, Jordan M. Winter, Myriam Gorospe, and Jonathan R. Brody
15. Cross-linking and Immunoprecipitation of Nuclear RNA Binding Proteins
Quan Li, Yuri Uemura, and Yukio Kawahara
16. Purifications of Non-coding RNA and Bound Proteins using FLAG Peptide-conjugated Antisense-oligonucleotides
Shungo Adachi and Tohru Natsume
Part III: Cell and Molecular Biological Approach Towards the Analysis of Noncoding RNAs and Nuclear Bodies
17. MMCT-Mediated Chromosome Engineering Technique Applicable to Functional Analysis of lncRNA and Nuclear Dynamics
Makiko Meguro-Horike and Shin-ichi Horike
18. Reconstitution of Nucleocytoplasmic Transport Using Digitonin-Permeabilized Cells
Shingo Kose, Tomoko Funakoshi, and Naoko Imamoto
19. Genome-wide Analysis of Long Noncoding RNA Turnover
Hidenori Tani, Naoto Imamachi, Rena Mizutani, Katsutoshi Imamura, Yeondae Kwon, Satoru Miyazaki, Sho Maekawa, Yutaka Suzuki, and Nobuyoshi Akimitsu
20. Knockdown of Nuclear-retained Long Noncoding RNAs Using Modified DNA Antisense Oligonucleotides
Xinying Zong, Lulu Huang, Vidisha Tripathi, Raechel Peralta, Susan M. Freier, Shuling Guo, and Kannanganattu V. Prasanth
21. siRNA Screening of Nuclear Proteins
Yuko Hasegawa and Shinichi Nakagawa
This volume focuses on cytological, biochemical, and molecular biological methods to identify and examine the function of each nuclear body, with an emphasis on the analysis of long non-coding RNAs. Chapters focus on exploring recent studies that reveal how certain long non protein-coding RNAs accumulate in specific nuclear bodies and regulate the function of the bodies by serving as architectural components or controlling the dynamics of associating protein components. Written in the highly successful Methods of 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 key tips on troubleshooting and avoiding known pitfalls.
Authoritative and practical, Nuclear Bodies and Noncoding RNAs: Methods and Protocols serves as a guideline for further study into the sophisticated regulation of gene expression.
Visualization of lncRNA by Single Molecule Fluorescence in situ Hybridization.- Super-resolution Imaging of Nuclear Bodies by STED Microscopy.- High-resolution 3D DNA FISH Using Plasmid Probes and Computational Correction of Optical Aberrations to Study Chromatin Structure at the Sub-megabase Scale.- Time-lapse Imaging of Nuclear Bodies.- Visualization of Nucleic Acids with Synthetic Exciton-controlled Fluorescent Oligonucleotide Probes.- Live CLEM Imaging to Analyze Nuclear Structures at High-resolution.- Ultrastructural Analysis of Nuclear Bodies Using Electron Microscopy.- Analyses of Nuclear Proteins and Nucleic Acid Structures Using Atomic Force Microscopy.- Genome-wide Co-localization Screening of Nuclear Body Components Using a Fluorescently Tagged FLJ cDNA Clone Library.- Purification of Specific Chromatin Regions Using Oligonucleotides: Capture Hybridization Analysis of RNA Targets (CHART).- RNA Antisense Purification (RAP) for Mapping RNA Interactions with Chromatin.- In situ Dissection of RNA Functional Subunits by Domain-specific Chromatin Isolation by RNA Purification (dChIRP).- Extracting, Enriching and Identifying Nuclear Body Sub-complexes Using Label-based Quantitative Mass Spectrometry.- Studying RNA-binding Protein Interactions with Target mRNAs in Eukaryotic Cells: Native Ribonucleoprotein Immunoprecipitation (RIP) Assays.- Cross-linking and Immunoprecipitation of Nuclear RNA Binding Proteins.- Purifications of Non-coding RNA and Bound Proteins using FLAG Peptide-conjugated Antisense-oligonucleotides.- MMCT-Mediated Chromosome Engineering Technique Applicable to Functional Analysis of lncRNA and Nuclear Dynamics.- Reconstitution of Nucleocytoplasmic Transport Using Digitonin-Permeabilized Cells.- Genome-wide Analysis of Long Noncoding RNA Turnover.- Knockdown of Nuclear-retained Long Noncoding RNAs Using Modified DNA Antisense Oligonucleotides.- siRNA Screening of Nuclear Proteins.
Inhaltsverzeichnis
Part I: Imaging of Noncoding RNAs and Nuclear Bodies
1. Visualization of lncRNA by Single Molecule Fluorescence in situ Hybridization
Margaret Dunagin, Moran N. Cabili, John Rinn, and Arjun Raj
2. Super-resolution Imaging of Nuclear Bodies by STED Microscopy
Yasushi Okada and Shinichi Nakagawa
3. High-resolution 3D DNA FISH Using Plasmid Probes and Computational Correction of Optical Aberrations to Study Chromatin Structure at the Sub-megabase Scale
Luca Giorgetti, Tristan Piolot, and Edith Heard
4. Time-lapse Imaging of Nuclear Bodies
Saskia Hutten, Samuel Swift, and Angus I. Lamond
5. Visualization of Nucleic Acids with Synthetic Exciton-controlled Fluorescent Oligonucleotide Probes
Dan Ohtan Wang and Akimitsu Okamoto
6. Live CLEM Imaging to Analyze Nuclear Structures at High-resolution
Tokuko Haraguchi, Hiroko Osakada, and Takako Koujin
7. Ultrastructural Analysis of Nuclear Bodies Using Electron Microscopy
Sylvie Souquere and Gérard Pierron
8. Analyses of Nuclear Proteins and Nucleic Acid Structures Using Atomic Force Microscopy
Jamie L. Gilmore, Aiko Yoshida, Hirohide Takahashi, Katashi Deguchi, Toshiro Kobori, Emilie Louvet, Masahiro Kumeta, Shige H. Yoshimura, and Kunio Takeyasu
9. Genome-wide Co-localization Screening of Nuclear Body Components Using a Fluorescently Tagged FLJ cDNA Clone Library
Tetsuro Hirose and Naoki Goshima
Part II: Biochemical Analysis of Noncoding RNAs and Nuclear Bodies
10. Purification of Specific Chromatin Regions Using Oligonucleotides: Capture Hybridization Analysis of RNA Targets (CHART)
Christopher P. Davis and Jason A. West
11. RNA Antisense Purification (RAP) for Mapping RNA Interactions with Chromatin
Jesse Engreitz, Eric S. Lander, and Mitchell Guttman
12. In situ Dissection of RNA Functional Subunits by Domain-specific Chromatin Isolation by RNA Purification (dChIRP)
Jeffrey J. Quinn and Howard Y. Chang
13. Extracting, Enriching and Identifying Nuclear Body Sub-complexes Using Label-based Quantitative Mass Spectrometry
Archa Fox, Virja Mehta, Severine Boulon, and Laura Trinkle-Mulcahy
14. Studying RNA-binding Protein Interactions with Target mRNAs in Eukaryotic Cells: Native Ribonucleoprotein Immunoprecipitation (RIP) Assays
Joseph A. Cozzitorto, Maya Jimbo, Saswati Chand, Fernando Blanco, Shruti Lal, Melissa Gilbert, Jordan M. Winter, Myriam Gorospe, and Jonathan R. Brody
15. Cross-linking and Immunoprecipitation of Nuclear RNA Binding Proteins
Quan Li, Yuri Uemura, and Yukio Kawahara
16. Purifications of Non-coding RNA and Bound Proteins using FLAG Peptide-conjugated Antisense-oligonucleotides
Shungo Adachi and Tohru Natsume
Part III: Cell and Molecular Biological Approach Towards the Analysis of Noncoding RNAs and Nuclear Bodies
17. MMCT-Mediated Chromosome Engineering Technique Applicable to Functional Analysis of lncRNA and Nuclear Dynamics
Makiko Meguro-Horike and Shin-ichi Horike
18. Reconstitution of Nucleocytoplasmic Transport Using Digitonin-Permeabilized Cells
Shingo Kose, Tomoko Funakoshi, and Naoko Imamoto
19. Genome-wide Analysis of Long Noncoding RNA Turnover
Hidenori Tani, Naoto Imamachi, Rena Mizutani, Katsutoshi Imamura, Yeondae Kwon, Satoru Miyazaki, Sho Maekawa, Yutaka Suzuki, and Nobuyoshi Akimitsu
20. Knockdown of Nuclear-retained Long Noncoding RNAs Using Modified DNA Antisense Oligonucleotides
Xinying Zong, Lulu Huang, Vidisha Tripathi, Raechel Peralta, Susan M. Freier, Shuling Guo, and Kannanganattu V. Prasanth
21. siRNA Screening of Nuclear Proteins
Yuko Hasegawa and Shinichi Nakagawa
Klappentext
This volume focuses on cytological, biochemical, and molecular biological methods to identify and examine the function of each nuclear body, with an emphasis on the analysis of long non-coding RNAs. Chapters focus on exploring recent studies that reveal how certain long non protein-coding RNAs accumulate in specific nuclear bodies and regulate the function of the bodies by serving as architectural components or controlling the dynamics of associating protein components. Written in the highly successful Methods of 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 key tips on troubleshooting and avoiding known pitfalls. nAuthoritative and practical, Nuclear Bodies and Noncoding RNAs: Methods and Protocols serves as a guideline for further study into the sophisticated regulation of gene expression.
Includes cutting-edge methods and protocols
Provides step-by-step detail essential for reproducible results
Contains key notes and implementation advice from the experts
Includes supplementary material: sn.pub/extras
