Bacterial Transcriptional Control, Softcover reprint of the original 1st ed. 2015 Methods and Protocols Methods in Molecular Biology Series, Vol. 1276

This volumeis designed to be a resource of proven techniques and approaches for probing the activities of bacterial, eukaryotic, and archaeal RNA polymerases. This book features a collection of in vitro and in vivo technologies that will permit researchers to purify and probe the position and stability of RNA polymerase complexes at different points of the transcription cycle, analyze the various translocations and intermolecular movements associated with catalysis, define recruitment strategies, probe the roles of transcription factors in each stage of the cycle, highlight conserved and disparate fidelity mechanisms, analyze the resultant transcripts, and study coordination of the nascent mRNA synthesis by the RNA polymerase and mRNA translation by the ribosome. 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 troubles troubleshooting and avoiding known pitfalls.

Practical and timely, Bacterial Transcriptional Controls: Methods and Protocols highlights the breadth and depth of techniques that are likely to continue shaping the transcription community in the future.

1. Mapping the Escherichia coli Transcription Elongation Complex with Exonuclease III
Zhaokun Liu and Irina Artsimovitch

2. Purification of Bacterial RNA Polymerase: Tools and Protocols
Vladimir Svetlov and Irina Artsimovitch

3. Monitoring Translocation of Multisubunit RNA Polymerase Along the DNA with Fluorescent Base Analogues
Anssi M. Malinen, Matti Turtola, and Georgiy A. Belogurov

4. In vitro and in vivo Methodologies for Studying the Sigma54-dependent Transcription
Martin Buck, Christoph Engl, Nicolas Joly, Goran Jovanovic, Milija Jovanovic, Edward Lawton, Christopher McDonald, Jörg Schumacher, Christopher Waite, and Nan Zhang

5. Methods for the Assembly and Analysis of in vitro Transcription-coupled-to-translation Systems
Daniel Castro-Roa and Nikolay Zenkin

6. Site-specific Incorporation of Probes into RNA Polymerase by Unnatural-amino-acid Mutagenesis and Staudinger-Bertozzi Ligation
Anirban Chakraborty, Abhishek Mazumder, Miaoxin Lin, Adam Hasemeyer, Qumiao Xu, Dongye Wang, Yon W. Ebright, and Richard H. Ebright

7. Reconstitution of Factor-dependent, Promoter Proximal Pausing in Drosophila Nuclear Extracts
Jian Li and David S. Gilmour

8. Direct Competition Assay for Transcription Fidelity
Lucyna Lubkowska and Maria L. Kireeva

9. Single-stranded DNA Aptamers for Functional Probing of Bacterial RNA Polymerase
Danil Pupov and Andrey Kulbachinskiy  

10. Biochemical Analysis of Transcription Termination by RNA Polymerase III from Yeast Saccharomyces cerevisiae
Aneeshkumar G. Arimbasseri and Richard J. Maraia

11. Use of RNA Polymerase Molecular Beacon Assay to Measure RNA Polymerase Interactions with Model Promoter Fragments
Vladimir Mekler and Konstantin Severinov

12. Preparation of cDNA Libraries for High-throughput RNA Sequencing Analysis of RNA 5’ Ends
Irina O. Vvedenskaya, Seth R. Goldman, and Bryce E. Nickels

13. In situ Footprinting of E. coli Transcription Elongation Complex with Chloroacetaldehyde
A. Rachid Rahmouni and Christine Mosrin-Huaman

14. Using Solutes and Kinetics to Probe Large Conformational Changes in the Steps of Transcription Initiation
Emily Ruff, Wayne S. Kontur, and M. Thomas Record Jr.

15. Manipulating Archaeal Systems to Permit Analyses of Transcription Elongation-termination Decisions in vitro
Alexandra M. Gehring and Thomas J. Santangelo

16. Purification of Active RNA Polymerase I from Yeast
Francis Dean Appling and David Alan Schneider

17. Transcription in Archaea: preparation of Methanocaldococcus jannaschii Transcription Machinery
Katherine Smollett, Fabian Blombach, and Finn Werner

18. Transcription in Archaea: in vitro Transcription Assays for mjRNAP
Katherine Smollett, Fabian Blombach, and Finn Werner

19. Experimental Analysis of hFACT Action during Pol II Transcription in vitro
Fu-Kai Hsieh, Olga I. Kulaeva, and Vasily M. Studitsky

20. ChIP-seq for Genome-scale Analysis of Bacterial DNA-binding Proteins
Richard P. Bonocora and Joseph T. Wade

Includes cutting-edge methods and protocols

Provides step-by-step detail essential for reproducible results

Contains key notes and implementation advice from the experts