Bacterial Regulatory RNA Methods and Protocols

The discovery of wide-spread RNA-based regulation in bacteria has led to new evaluations of the importance of bacterial regulatory RNA in every aspect of bacterial physiology. In Bacteria Regulatory RNA: Methods and Protocols, expert researchers in the field detail many of the methods which are now commonly used to study bacterial regulatory RNA. These include methods and techniques to identify regulatory RNAs, characterizing the function and expression of regulatory RNAs in bacterial cells, RNA structure prediction, and interactions between regulatory RNAs and proteins. 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.

 Authoritative and practical, Bacteria Regulatory RNA: Methods and Protocols seeks to aid scientists in the further study of bacterial regulatory RNA.

Part I- Identification of Regulatory RNAs

1. Bioinformatic Discovery of Bacterial Regulatory RNAs using SIPH

Jonathan Livny

2. How to Identify CRISPRs in Sequencing Data

Christine Drevet and Christine Pourcel

3. A Strategy for Identifying Non-coding RNAs using Whole-genome Tiling Arrays

Stephen G. Landt and Eduardo Abeliuk

4. Genetic Screens to Identify Bacterial sRNA Regulators

Pierre Mandin

Part II - Function and Expression of Regulatory RNAs

5. Competition Assays Using Barcoded Deletion Strains to Gain Insight Into Small RNA Function

Errett C. Hobbs and Gisela Storz

6. Use of Semi-quantitative Northern Blot Analysis to Determine Relative Quantities of Bacterial CRISPR Transcripts

Ksenia Pougach and Konstantin Severinov

7. RNA Visualization in Bacteria by Fluorescence in Situ Hybridization

Jay H. Russell and Kenneth C. Keiler

Part III - RNA Structure and Biophysical Measurements

 8. RNA Structure Prediction: An Overview of Methods

Matthew G. Seetin and David H. Mathews

9. Crystallization of RNA-Protein Complexes: From Synthesis and Purification of Individual Components to Crystals

Anna Perederina and Andrey S. Krasilnikov

10. Analysis of RNA Folding and Ligand Binding by Conventional and High-Throughput Calorimetry

Joshua E. Sokoloski and Philip C. Bevilacqua

Part IV – RNA-Protein Interactions

11. Use of Aptamer Tagging to Identify in vivo Protein Binding Partners of Small Regulatory RNAs

Colin P. Corcoran,  Renate Rieder, Dimitri Podkaminski, Benjamin Hofmann, and Jörg Vogel

12. Gel Mobility Shift Assays to Detect Protein-RNA Interactions

Alexander V. Yakhnin, Helen Yakhnin and Paul Babitzke

13. RNase Footprinting of Protein Binding Sites on an mRNA Target of Small RNAs

Peng Yi, Soper, Toby J., and Woodson Sarah A

Part V – RNA-RNA Interactions

14.Computational Identification of sRNA Targets

Brian Tjaden

 15. Detection of sRNA-mRNA Interactions by Electrophoretic Mobility Shift Assay

Teppei Morita , Kimika Maki, and Hiroji Aiba

16. Activity of Small RNAs on the Stability of Targeted mRNAs in vivo

Guillaume Desnoyers and Eric Massé

Part VI –Interactions of Regulatory RNA with RNA Polymerase and the Ribosome

17. Native Gel Electrophoresis to Study the Binding and Release of RNA Polymerase by 6S RNA

Karen M. Wassarman

18. Ribosome Purification Approaches for Studying Interactions of Regulatory Proteins and RNAs with the Ribosome

Preeti Mehta, Perry Woo, Krithika Venkataraman, and A. Wali Karzai

19. Analysis of Aminoacyl- and Peptidyl-tRNAs by Gel Electrophoresis

Brian D. Jansse, Elie J. Diner, and Christopher S. Hayes

20. In vitro trans-translation Assays

Daisuke Kurit, Akira Muto, and Hyouta Himeno