exploring genome-wide organization of chromatin structure...
TRANSCRIPT
Exploring Genome-wide
Organization of Chromatin
Structure by ChIP
Alon Goren, Ph.D.
Broad Institute of MIT & Harvard,
MGH Pathology Harvard Medical School
Webinar Overview
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Introduction to chromatin, ChIP (Chromatin IP) and uses of the method
Overview of genomic approaches to map in-vivo chromatin structure
Detailed description of genome-wide chromatin organization
Usage of genome visualization tools
Miriam Ferrer, Abcam speaker
(highlighting ChIP grade antibodies, ChIP kits)
Questions and answers from delegates
Major scientific discoveries stemming from charting of
in-vivo chromatin maps
Part I
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Introduction to chromatin, ChIP (Chromatin IP) and uses of the method
Overview of genomic approaches to map in-vivo chromatin structure
Introduction to Chromatin Structure & Organization
Source: Ecker et al., Nature 2012
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Introduction to Chromatin Structure & Organization
Source: Zhou*, Goren* & Bernstein Nature Reviews Genetics 2010
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Layers of chromatin
organization
Introduction to ChIP – Chromatin ImmunoPrecipitation
Map the organization of histone modifications (HMs),
transcription factors (TFs) and chromatin regulators (CRs) Goal
ChIP
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Overview of ChIP-chip & ChIP-seq
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Source: Schones & Zhao, Nature Reviews Genetics 2008
ChIP-chip ChIP-seq
Overview of ChIP-chip & ChIP-seq
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Source: Schones & Zhao, Nature Reviews Genetics 2008
ChIP-seq ChIP-chip
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Source: Park Nature Reviews Genetics 2009
Comparison of ChIP-chip and ChIP-seq
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ChIP-seq – a Peak Into the Computational Approach
Source: Mikkelsen et al., Nature 2007
A. Align reads
B. Infer positions of ChIP fragments
C. Count fragments at each genomic position
Computational pipeline
Sequencing
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Approaches for the Analyses of ChIP-seq Data
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Source: Park Nature Reviews Genetics 2009
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(WCE)
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Part II
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Detailed description of genome-wide chromatin organization
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Source: Ernst et al., Nature 2011
Chromatin States – Discrete Genomic Combinations of HMs
Chromatin States – Discrete Genomic Combinations of HMs
Source: Barth et al., Trends in Bio Sci. 2010
TSS TTS Gene body
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Five Chromatin Types in Drosophila Cells
Source: Filion et al., Cell 2010
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A Model – Fine-Tuning of Genomic Elements by Histone Modifications
Source: Zhou*, Goren* & Bernstein Nature Reviews Genetics 2010
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The Complexity of CRs
>100 Isozymes – HDACs, HDMs, HMTs, HATs
Tissue-specific, developmentally-regulated
Amplified, mutated, rearranged in cancer
Source: Kouzarides, Cell 2007
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Part III
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Usage of genome visualization tools
Genomic Visualization and Analysis Tools: 3. UCSC:
Source: http://genome.ucsc.edu/
More at http://www.slideshare.net/joachimjacob/bits-ucsc-genome-browser
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Genomic Visualization Tools – IGV
Source: http://www.broadinstitute.org/igv/
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Genomic Visualization Tools – IGV
Source: Jim Robinson, The Broad Institute
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Genomic Visualization Tools – IGV
Source: Jim Robinson, The Broad Institute
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Genomic Analysis Tools: Genomespace
Source: http://www.genomespace.org/
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Genomespace Principles
Source: http://www.genomespace.org/ http://www.slideshare.net/jandot/m-reich-genomespace
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Genomic Visualization and Analysis Tools: 1. Genomespace
Source: http://www.genomespace.org/ http://www.slideshare.net/jandot/m-reich-genomespace
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Part IV
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Major scientific discoveries stemming from charting of
in-vivo chromatin maps
Enhancers Identification and Characterization
Sources: Heintzman*, Hon* & Hawkins* et al., Nature 2009; Shen*, Yue.* et al., Nature 2012
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Introduction to Chromatin Regulators (CRs)
H3K36me3 H3K4me3 H3K27me3
CRs
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Combinatorial Patterning of Chromatin Regulators (CRs)
Source: Ram*, Goren* et al. Cell 2011
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CRome web portal
Source: www.broadinstitute.org/software/CRome
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ENCODE Consortium Studies
Source: http://www.nature.com/encode/ Find more at http://www.slideshare.net/mateongenaert/encode-project-brief-summary-of-main-findings
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Identification and Characterization Regulatory Regions
Source: Neph*, Vierstra*, Stergachis*, Reynolds* et al., Nature 2012
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Dynamic profiling of TF binding by High-throughput ChIP-seq
Source: Garber*, Yosef*, Goren et al., Molecular Cell 2012
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Future Directions: Developmental Epigenomics
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Acknowledgments
Melissa Gymrek
Jeff Xing
Daniel Fernandez
Timothy Durham
Oren Ram
Noam Shoresh
Chuck Epstein
Broad Epigenomics
Lesley Gafney
Dongkeun Jang
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Janis Lem
Alex Meissner
Ido Amit
Nir Yosef
Jason Ernst
Jim Robinson
Eric Mendenhall
Leah Escalante
Brad Bernstein
Aviv Regev
ENCODE consortium
Charles H. Hood Foundation
Post-doctoral Fellowship
Centers of Excellence in
Genomic Science (CEGS)
Part V
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Miriam Ferrer, Abcam speaker
(highlighting ChIP grade antibodies, ChIP kits)
Protocol and troubleshooting guidebook
2nd Edition of our general protocol
and troubleshooting book
• ChIP
• IHC
• Flow Cytometry
• …and many more
Want a copy?
• Contact:
Mention ‘Protocol book’
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Protocol and troubleshooting guidebook
• www.abcam.com/epigenetics
• www.abcam.com/protocols
CLIP protocol
• www.abcam.com/CLIPprotocol
RIP protocol
• www.abcam.com/RIPprotocol
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How to contact us
US
• Email: [email protected]
• Tel: +1 888-77-ABCAM (22226)
or 617-225-2272
• Website: www.abcam.com
Hong Kong
• Email: [email protected]
• Tel: +(852) 2603 6823
• Website: www.abcam.cn
UK
• Email: [email protected]
• Tel: +44 (0) 1223 696000
• Website: www.abcam.com
Japan
• Email: [email protected]
• Tel: +81 (0) 3 6231 0940
• Website: www.abcam.co.jp
Abcam’s Scientific Support Team
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www.abcam.com/epigenetics
Abcam’s new online tool for
epigenetics
• The hottest products
• The newest protocols
• The best meetings
• In one easy location!
Bulk up and save
• Buying in bulk allows you to…
• Save money
• Minimize variability
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EpiSeeker ChIP Kits
Your tool for advancing epigenetics
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Available EpiSeeker ChIP kits
General
• One Step
• Plant
• MBD2(Methyl-CpG Binding Domain
protein 2)
Methylated histone modifications
Optimized for cells or tissue
• Histone H3 (methyl K4)
• Histone H3 (methyl K9) &
(tri-methyl K9)
• Histone H3 (methyl K27)
Acetylated histone modifications
Optimized for cells or tissue
• Histone H3 (acetyl)
• Histone H4 (acetyl)
Methylated DNA
• MeDIP – 5mC (optimized for cells,
tissue or nuclear extracts)
• hMeDIP – 5hmC
(All EpiSeeker kits are X-ChIP)
Resources
• http://www.abcam.com/episeeker
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Characteristics EpiSeeker ChIP kits
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Simple to
use
Reaction takes place in the wells of the 96-wp: easy for
standardization and HTP
Quick Only 5 hours to perform ChIP (compared to 2 days using
conventional method)
Inclusive Kit contains all necessary reagents for ChIP reaction (excluding
cross-linking step)
Ready to go Pre-selected ChIP-grade antibody: optimized for assay
Eluted DNA can be processed straight away by any DNA-
amplification method (ChIP-ChIP, ChIP-seq) Compatible
EpiSeeker MeDIP Kits
• Specific antibody to enrich 5-mC/5-
hmC rich DNA regions
• MeDIP –detects 5-mC
(methylated cytosine) residues
• hMEDIP – detects 5-hmC
(hydroxymethylated cytosine). 5-hmC
is a recently discovered modification
in animal cells apparently involved in
DNA demethylation, but its role is not
clear
• Ready in less than 4 hours
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Webinar attendee promotion
35% Off ChIP Kits
Webinar attendees will receive a discount
code for 35% off the following products
• Ab500 (standard ChIP kit)
• Any ChIP Kit on the EpiSeeker range
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Chromatin, Replication and Chromosomal Stability 2013
Date: June 17-19, 2013
Location: Copenhagen, Denmark
Conference Topics
• Chromosomal architecture
• Chromosomal stability
• Replication structure and fork progression
• Chromatin maintenance and cellular
memory
Keynote speakers
• Susan Gasser and Adrian Bird
Share your research
• Talk and poster places opportunities
available
Meeting website:
• www.abcam.com/copenhagen2013
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Questions?
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