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Circadian Rhythms and Biological Clocks Part A
1st Edition, Volume 551 - January 27, 2015
Editor: Amita Sehgal
Language: English
Hardback ISBN:9780128012185
9 7 8 - 0 - 1 2 - 8 0 1 2 1 8 - 5
eBook ISBN:9780128013410
9 7 8 - 0 - 1 2 - 8 0 1 3 4 1 - 0
Two new volumes of Methods in Enzymology continue the legacy of this premier serial with quality chapters authored by leaders in the field. Circadian Rhythms and Biologica…Read more
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Two new volumes of Methods in Enzymology continue the legacy of this premier serial with quality chapters authored by leaders in the field. Circadian Rhythms and Biological Clocks Part A and Part B is an exceptional resource for anybody interested in the general area of circadian rhythms. As key elements of timekeeping are conserved in organisms across the phylogenetic tree, and our understanding of circadian biology has benefited tremendously from work done in many species, the volume provides a wide range of assays for different biological systems. Protocols are provided to assess clock function, entrainment of the clock to stimuli such as light and food, and output rhythms of behavior and physiology. This volume also delves into the impact of circadian disruption on human health. Contributions are from leaders in the field who have made major discoveries using the methods presented here.
Continues the legacy of this premier serial with quality chapters authored by leaders in the field
Covers research methods in biomineralization science
Keeping with the interdisciplinary nature of the circadian rhythm field, the volume includes diverse approaches towards the study of rhythms, from assays of biochemical reactions in unicellular organisms to monitoring of behavior in humans.
Biochemists, biophysicists, molecular biologists, analytical chemists, and physiologists.
Preface
Part I: Organismal Rhythms as Read-Outs of Clock Function
Chapter One: Studying Circadian Rhythm and Sleep Using Genetic Screens in Drosophila
Abstract
1 Introduction: Studying Circadian Behavior in the Fruit Fly, Drosophila melanogaster
2 Screening for Circadian Rhythm and Sleep Mutants
3 Screening Techniques
Acknowledgments
Chapter Two: Dissecting the Mechanisms of the Clock in Neurospora
Abstract
1 Introduction
2 Molecular Mechanism of the Neurospora Circadian Oscillator
3 Core Clock Components
4 Conclusion
Chapter Three: High-Throughput and Quantitative Approaches for Measuring Circadian Rhythms in Cyanobacteria Using Bioluminescence
Abstract
1 Theory
2 Build a Computer-Controlled Turntable
3 Use a Computer-Controlled Turntable
4 Analyzing Data from Turntable
5 Steps to Extract Reliable Quantitative Information from Bioluminescence Levels
Acknowledgments
Chapter Four: Using Circadian Entrainment to Find Cryptic Clocks
Abstract
1 Introduction
2 Methods
3 Discussion
Acknowledgments
Chapter Five: Wavelet-Based Analysis of Circadian Behavioral Rhythms
Abstract
1 Introduction
2 Fourier and Wavelet Methods for Time Series Analysis
3 Computations
4 Concluding Remarks
Chapter Six: Genetic Analysis of Drosophila Circadian Behavior in Seminatural Conditions
Abstract
1 Introduction
2 Considerations for Studies Outside
3 Simulating Natural Conditions in the Laboratory
Acknowledgments
Part II: Characterization of Molecular Clock Components
Chapter Seven: Methods to Study Molecular Mechanisms of the Neurospora Circadian Clock
Abstract
1 Introduction
2 Description of Methods
3 Concluding Remarks
Acknowledgment
Chapter Eight: Detecting KaiC Phosphorylation Rhythms of the Cyanobacterial Circadian Oscillator In Vitro and In Vivo
Abstract
1 Theory
2 Equipment
3 Materials
4 Protocol
5 Step 1: Expression of KaiA or KaiB in E. coli
6 Step 2: Expression of KaiC in E. coli
7 Step 3: Purification of KaiA or KaiB
8 Step 4: Purification of KaiC
9 Step 5: In vitro oscillation reaction
10 Step 6: SDS-PAGE
11 Step 7: Densitometry
12 Detection of protein phosphorylation forms from in vivo cell extracts
13 Equipment
14 Materials
15 Protocol
16 Step 1: Preparation
17 Step 2: Electrophoresis and Blotting
Acknowledgments
Chapter Nine: The Role of Casein Kinase I in the Drosophila Circadian Clock
Abstract
1 Introduction
2 Expression of Mutant Forms of DBT with the GAL4/UAS Binary Expression Method
3 Expression of DBT in Drosophila S2 Cells for Analysis of DBT Kinase Activity
4 Proteomic Approaches
Chapter Ten: Purification and Analysis of PERIOD Protein Complexes of the Mammalian Circadian Clock
Abstract
1 General Strategy
2 Extraction and Characterization of PER Complexes from Mouse Tissues
Chapter Eleven: Best Practices for Fluorescence Microscopy of the Cyanobacterial Circadian Clock
Abstract
1 Introduction
2 Materials
3 Methods
Acknowledgments
Chapter Twelve: Structural and Biophysical Methods to Analyze Clock Function and Mechanism
Abstract
1 Introduction
2 Kai Protein Overexpression, Purification, Complex Formation, and Analysis by Denatured and Native Polyacrylamide Gel Electrophoresis
3 Analytical Ultracentrifugation
4 Dynamic Light Scattering
5 Thin Layer Chromatography
6 Mass Spectrometry
7 Site-Directed Mutagenesis
8 Fluorescence Techniques (Labeled Proteins, Anisotropy, and Fluorescence Resonance Energy Transfer)
9 Electron Microscopy
10 X-ray Crystallography
11 Small-Angle X-ray and Neutron Scattering
12 Nuclear Magnetic Resonance
13 Hydrogen–Deuterium Exchange
14 MD Simulations
15 Modeling the In Vitro Oscillator
16 Summary and Outlook
Acknowledgments
Chapter Thirteen: Identification of Small-Molecule Modulators of the Circadian Clock
Abstract
1 Introduction
2 Cell-Based Circadian Assay
3 High-Throughput Screening System
4 Circadian Screening
5 Conclusion
Part III: Circadian Regulation of Gene and Protein Expression
Chapter Fourteen: ChIP-seq and RNA-seq Methods to Study Circadian Control of Transcription in Mammals
Abstract
1 Critical Factors
2 ChIP-seq Method for Mouse Liver
3 RNA-Seq Method for Mouse Liver
Chapter Fifteen: ChIPping Away at the Drosophila Clock
Abstract
1 Introduction
2 Equipment
3 Solutions
4 Protocol
5 Discussion
Chapter Sixteen: Considerations for RNA-seq Analysis of Circadian Rhythms
Abstract
1 Introduction
2 Results
3 Conclusions
4 Methods
Acknowledgments
Chapter Seventeen: RNA-seq Profiling of Small Numbers of Drosophila Neurons
Abstract
1 Introduction
2 Results/Methods
3 Discussion
Chapter Eighteen: Analysis of Circadian Regulation of Poly(A)-Tail Length
Abstract
1 Introduction
2 Measurement of Poly(A)-Tail Length at a Genomewide Level
3 Measurement of Poly(A)-Tail Length at a Single-Gene Level
4 Materials
5 Concluding Remarks
Acknowledgments
Chapter Nineteen: Sample Preparation for Phosphoproteomic Analysis of Circadian Time Series in Arabidopsis thaliana
Abstract
1 Introduction
2 Materials and Methods
3 Results
4 Discussion
5 Conclusions
Acknowledgments
Author Index
Subject Index
No. of pages: 488
Language: English
Edition: 1
Volume: 551
Published: January 27, 2015
Imprint: Academic Press
Hardback ISBN: 9780128012185
eBook ISBN: 9780128013410
AS
Amita Sehgal
Dr. Sehgal is the John Herr Musser Professor of Neuroscience and Director of a Program in Chronobiology at the Perelman School of Medicine, University of Pennsylvania. She has also been an Investigator of the Howard Hughes Medical Institute since 1997. Dr. Sehgal received her Ph.D. from the Graduate School of Medical Sciences, Cornell University, working with Dr. Moses Chao, and conducted her postdoctoral work with Dr. Michael Young at Rockefeller University. A major focus in her laboratory is the cellular and molecular basis of circadian rhythms, using the fruit fly, Drosophila melanogaster as a model system. Her laboratory also developed a Drosophila model for the study of sleep, which has been adopted by laboratories worldwide, and is rapidly provding insight into the regulation and function of sleep. In 2009, Dr. Sehgal was elected to the Institute of Medicine and in 2011 to the American Academy of Arts and Sciences.
Affiliations and expertise
John Herr Musser Professor of Neuroscience; co-Director, Penn Medicine Neuroscience Center, Perelman School of Medicine, University of Pennsylvania, Philadelphia, USA
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