Long Non-Coding RNAs in Cancer

This volume presents techniques needed for the study of long non-coding RNAs (lncRNAs) in cancer from their identification to functional characterization. Chapters guide readers through identification of lncRNA expression signatures in cancer tissue or liquid biopsies by RNAseq, single Cell RNAseq, Phospho RNAseq or Nanopore Sequencing techniques; validation of lncRNA signatures by Real time PCR, digital PCR or in situ hybridization; and functional analysis by siRNA or CRISPR based methods for lncRNA silencing or overexpression. Lipid based nanoparticles for delivery of siRNAs in vivo, lncRNA-protein interactions, viral lncRNAs and circRNAs are also treated in this volume. 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 practical, Long Non-Coding RNAs in Cancer aims to provide a collection of laboratory protocols, bioinformatic pipelines, and review chapters to further research in this vital field.

Includes cutting-edge methods and protocols

Provides step-by-step detail essential for reproducible results

Contains key notes and implementation advice from the experts

Klappentext

This volume presents techniques needed for the study of long non-coding RNAs (lncRNAs) in cancer from their identification to functional characterization. Chapters guide readers through identification of lncRNA expression signatures in cancer tissue or liquid biopsies by RNAseq, single Cell RNAseq, Phospho RNAseq or Nanopore Sequencing techniques; validation of lncRNA signatures by Real time PCR, digital PCR or in situ hybridization; and functional analysis by siRNA or CRISPR based methods for lncRNA silencing or overexpression. Lipid based nanoparticles for delivery of siRNAs in vivo, lncRNA-protein interactions, viral lncRNAs and circRNAs are also treated in this volume. 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 practical, Long Non-Coding RNAs in Cancer aims to provide a collection of laboratory protocols, bioinformatic pipelines, and review chapters to further research in this vital field.

Inhalt

PART I: LNCRNA CLASSIFICATION AND UTILITY IN CANCER

1. LncRNAs and Available Databases

   Sara Napoli

2. LncRNA as Cancer Biomarkers

   Ingram Iaccarino and Wolfram Klapper

   PART II: LNCRNA PROFILING AND DISCOVERY

   3. Bioinformatics Pipeline to Analyze lncRNA Arrays

   Katia Todoerti, Domenica Ronchetti, Martina Manzoni, Elisa Taiana, Antonino Neri, and Luca Agnelli

3. Bioinformatic Pipelines to Analyze lncRNAs RNAseq Data

   Luca Agnelli, Stefania Bortoluzzi, and Giancarlo Pruneri

   5. Single Cell RNAseq Analysis of lncRNAs Stefano Cagnin, Enrico Alessio, Raphael Severino Bonadio, and Gabriele Sales

   PART III: VALIDATION OF IDENTIFIED LNCRNAS

4. Measuring lncRNA Expression by Real Time PCR

   Sonam Dhamija and Manoj B. Menon

5. Quantitation of Long Non-coding RNA using Digital PCR

   Yu Ota, Irene K Yan, and Tushar Patel

6. Detection of lncRNA by LNA-based In Situ Hybridization in Paraffin Embedded Cancer Cell Spheroids

   Boye Schnack Nielsen, Jesper Larsen, Jakob Høffding, Son Ly Nhat, Natasha Helleberg Madsen, Trine Møller, Bjørn Holst, Kim Holmstrøm

   PART IV: FUNCTIONAL STUDIES TO DECIPHER THE ROLE OF IDENTIFIED LNCRNAS

7. In vitro Silencing of lncRNA Expression using siRNAs Meike Thijssen, Jennifer Bintz, and Luis Arnes

8. In vitro silencing of lncRNAs using LNA gapmeRs

   Elisa Taiana, Vanessa Favasuli, Domenica Ronchetti, Eugenio Morelli, Pierfrancesco Tassone, Giuseppe Viglietto, Nikhil C. Munshi, Antonino Neri, and Nicola Amodio

9. Methods used to make Lipid Nanoparticles to deliver LNA Gapmers against lncRNAs into Acute Myeloid Leukemia (AML) Blasts Chun-Tien Kuo, Robert Lee, and Ramiro Garzon

10. CRISPR/Cas9 to Silence Long Non-coding RNAs

    Ingrid Arctander Rosenlund, George A. Calin, Mihnea P. Dragomir,and Erik Knutsen

11. CRISPR interference (CRISPRi) and CRISPR activation (CRISPRa) to explore the oncogenic lncRNA network

    Eugenio Morelli, Annamaria Gulla', Nicola Amodio, Elisa Taiana, Antonino Neri, Mariateresa Fulciniti, and Nikhil C. Munshi

    14. In vivo Silencing/Overexpression of lncRNAs by CRISPR/Cas System Marianna Vitiello, Laura Poliseno, and Pier Paolo Pandolfi

12. Experimental Validation of the Non-Coding Potential for lncRNAs

    Emily Dangelmaier and Ashish Lal

13. Identification of lncRNA-protein interactions by CLIP and RNA pull-down assays

    Kunming Zhao, Xingwen Wang, and Ying Hu

14. Empirical Validation of Overlapping Virus lncRNAs and Coding Transcripts by Northern Blot

    Mehmet Kara and Scott A. Tibbetts

    PART V: LNCRNA ANALYSIS IN LIQUID BIOPSY

15. Phospho-RNAseq profiling of extracellular mRNAs and lncRNAs

    Maria D. Giraldez and Muneesh Tewari

    19. Detection of Circulating RNA using Nanopore Sequencing Jennifer Lindemann, Irene K Yan, and Tushar Patel

    20. LncRNA Quantification from Extracellular Vesicles Isolated from Blood Plasma or Conditioned Media

    Joan J. Castellano, Jordi Canals, Bing Han, Tania Díaz, Mariano Monzo, and Alfons Navarro

    PART VI: CIRCULAR RNAS

    21. The use of circRNAs as Biomarkers of Cancer Carla Solé, Gartze Mentxaka, and Charles H. Lawrie

16. Bioinformatic Analysis of Circular RNA Expression

    Enrico Gaffo, Alessia Buratin, Anna Dal Molin, and Stefania Bortoluzzi

    23. Study of circular RNAs (circRNAs) expression Eleonora D'Ambra and Mariangela Morlando