Description
In Polyadenylation: Methods and Protocols, expert researchers in the field detail many of the protocols which are now commonly used to study polyadenylation. Focusing on recent advances in the fast-moving polyadenylation filed, that has recently been recognized as a key contributor to the complexity of mammalian gene expression. 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 key tips on troubleshooting and avoiding known pitfalls. PART I: SECTION I: 3′-END PROCESSING IN EUKARYOTES 1. 3′ end Formation and Regulation of Eukaryotic mRNAs Thomas J. Sweet and Donny D. Licatalosi PART II: I-A General Methods 2. Poly(A) Polymerase-based Poly(A) Length Assay Deepak P. Patil, Baskar Bakthavachalu, and Daniel R. Schoenberg 3. Using Klenow Mediated Extension to Measure Poly(A)-tail Length and Position in the Transcriptome Man C. Lee, Amrei Jnicke, and Traude H. Beilharz 4. Detection of Uridylated mRNAs Franois M. Sement, Dominique Gagliardi 5. Cytoplasmic Polyadenylation Assays Olga Coll, Ana Villalba, Ftima Gebauer 6. In vitro Analysis of Transcriptional Activators and Polyadenylation Takashi Nagaike and James L. Manley 7. Fluorescence Detection of Polyadenylation Reaction through the Coordination of Adensoine2-coralyne-adensoine2 Jia-Hui Lin and Wei-Lung Tseng 8. Small Molecule Recognition of Poly(A) zgl Persil etinkol 9. The Stem-loop Luciferase Assay for Polyadenylation (SLAP) Method for Determining CstF-64-dependent Polyadenylation Activity J. Andrew Hockert and Clinton C. MacDonald PART III: I-B Alternative Polyadenylation I-B-1 Prediction of Polyadenylation Sites 10. Accurate Mapping of Cleavage and Polyadenylation Sites by 3′ Region Extraction and Deep Sequencing Mainul Hoque, Wencheng Li, and Bin Tian 11. Polyadenylation Site Prediction using PolyA-iEP Method Ioannis Kavakiotis, George Tzanis, and Ioannis Vlahavas 12. Detecting Alternative Polyadenylation from Microarray Data Antonio Lembo and Paolo Provero Part IV: I-B-2 Large Scale/Genome Wide Analysis of mRNA Polyadenylation and Application of the Next Generation DNA Sequencing in Polyadenylation Study 13. Quantitative Polyadenylation Site Mapping with Single-molecule Direct RNA Sequencing Fatih Ozsolak 14. RHAPA: A New Method to Quantify Alternative Polyadenylation Ashley L. Cornett and Carol S. Lutz 15. Multiplex Analysis of PolyA-linked Sequences (MAPS): An RNA-seq Strategy to Profile poly(A+) RNA Yu Zhou, Hai-Ri Li, Jie Huang, Ge Jin, and Xiang-Dong Fu 16. Global and Quantitative Profiling of Polyadenylated RNAs using PAS-seq Chengguo Yao and Yongsheng Shi 17. High-throughput Sequencing of RNA Isolated by Crosslinking and Immunoprecipitation (HITS-CLIP) to Determine Sites of Binding of CstF-64 on Nascent RNAs Petar N. Grozdanov and Clinton C. MacDonald Part V: BACTERIA AND ORGANELLES 18. Polyadenylation in Bacteria and Organelles Joanna Rorbach, Agnieszka J. Bobrowicz, Sarah Pearce, and Michal Minczuk. 19. In vivo Analysis of Polyadenylation in Prokaryotes Bijoy K. Mohanty and Sidney R. Kushner 20. Polyadenylation of RNA in E. coli: RNA Polymerase-associated (rA)n-synthetic Activities Grant A. Encalade and Maxim V. Sukhodolets 21. Analysis of the Human Mitochondrial Transcriptome using Directional Deep sequencing and Parallel Analysis of RNA Ends Oliver Rackham and Aleksandra Filipovska 22. Measurement of Mitochondrial RNA Stability by Metabolic Labeling of Transcripts with 4-Thiouridine Lukasz S. Borowski and Roman J. Szczesny Part VI: DEADENYLATION AND DECAY 23. Deadenylation and its Regulation in Eukaryotic Cells Xiaokan Zhang, Frida E. Kleiman, and Emral Devany 24. Assaying mRNA Deadenylation in vitro Mandy Jeske, Claudia Temme, and Elmar Wahle 25. Assaying mRNA Deadenylation in vivo Claudia Temme and Elmar Wahle 26. In vitro Analysis of RNA Degradation Catalyzed by Deadenylase Enzymes Joel Hrit, Nathan Raynard, Jamie Van Etten, Kamya Sankar, Adam Petterson, and Aaron C. Goldstrohm 27. microRNA-mediated Deadenylation in a Mammalian Cell-free System Motoaki Wakiyama, Shigeyuki Yokoyama 28. Quantitative Analysis of Deadenylation-independent mRNA Decay by a Modified MBRACE Assay Julie A. Dougherty, Roshan Mascarenhas, and Daniel R. Schoenberg.
