Main description:

This detailed volume assembles a number of the most commonly used and state-of-the-art methodologies in the revolutionary field of mRNA processing in order to aid researchers in choosing the best method for their specific problems. Given that the vast majority of alternative mRNA processing events has yet to be explored, these techniques promise to breathe new life into the myriad new ideas currently underway. 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.
Practical and easy to use, mRNA Processing: Methods and Protocols seeks to aid scientists venturing into what's certain to be another explosive period of growth in this dynamic field of study.
The chapter 'Robust, Cost-Effective Profiling of RNA Binding Protein Targets with Single-end Enhanced Crosslinking and Immunoprecipitation (seCLIP)' is published open access under a CC BY 4.0 license at link.springer.com.

Contents:

1. Detection and Identification of Uncapped RNA by Ligation-Mediated Reverse Transcription Polymerase Chain Reaction

C. Kiong Ho

2. Preparation of Splicing Competent Nuclear Extract from Mammalian Cells and In Vitro Pre-mRNA Splicing Assay

Maliheh Movassat, Hossein Shenasa, and Klemens J. Hertel

3. Single-Molecule Analysis of Pre-mRNA Splicing with Colocalization Single-Molecule Spectroscopy (CoSMoS)

Joerg E. Braun and Victor Serebrov

4. In Vitro Modulation of Endogenous Alternative Splicing Using Splice-Switching Antisense Oligonucleotides

Jeong Eun Park and Luca Cartegni

5. Purification of mRNA Processing Complexes Using an RNA Affinity Approach

Xiuye Wang and Yongsheng Shi

6. Polyadenylation Site-Based Analysis of Transcript Expression by 3'READS+

Dinghai Zheng and Bin Tian

7. Comprehensive Identification of mRNA Polyadenylation Sites by PAPERCLIP

Hun-Way Hwang and Robert B. Darnell

8. Microinjection and Fluorescence In Situ Hybridization Assay for Studying mRNA Export in Mammalian Cells

Ke Wang, Min Shi, and Hong Cheng

9. Methods for the Detection of Adenosine-to-Inosine Editing Events in Cellular RNA

Eimile Oakes, Pranathi Vadlamani, and Heather A. Hundley

10. Using RNA-Seq to Discover Genetic Polymorphisms that Produce Hidden Splice Variants

Shayna Stein, Emad Bahrami-Samani, and Yi Xing

11. Inducible Expression of Eukaryotic Circular RNAs from Plasmids

Deirdre C. Tatomer, Dongming Liang, and Jeremy E. Wilusz

12. Methods for Extraction of RNA, Proteins, or Protein Complexes from Subcellular Compartments of Eukaryotic Cells

Kyu-hyeon Yeom and Andrey Damianov

13. Isolation of Newly Transcribed RNA Using the Metabolic Label 4-Thiouridine

Angela Garibaldi, Francisco Carranza, and Klemens J. Hertel

14. Robust, Cost-Effective Profiling of RNA Binding Protein Targets with Single-End Crosslinking and Immunoprecipitation (seCLIP)

Eric L. Van Nostrand, Thai B. Nguyen, Chelsea Gelboin-Burkhart, Ruth Wang, Steven M. Blue, Gabriel A. Pratt, Ashley L. Louie, and Gene W. Yeo

15. Purification of Transcript-Specific mRNP Complexes Formed In Vivo from Saccharomyces cerevisiae

Jenna E. Smith and Kristian E. Baker

16. A Cell-Based High-Throuput Method for Identifying Modulators of Alternative Splicing

Sika Zheng

17. Genome-Wide RNAi Screens for RNA Processing Events in Drosophila Melanogaster S2 Cells

Natoya J. Peart and Eric J. Wagner

18. Assaying RNA Structure Inside Living Cells with SHAPE

Chao Feng, Dalen Chan, and Robert C. Spitale