exploring genome-wide organization of chromatin structure...

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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Viewing ChIP-seq tracks on UCSC

Source: http://genome.ucsc.edu/

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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:

mailrequest@abcam.com

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: us.technical@abcam.com

• Tel: +1 888-77-ABCAM (22226)

or 617-225-2272

• Website: www.abcam.com

Hong Kong

• Email: hk.technical@abcam.com

• Tel: +(852) 2603 6823

• Website: www.abcam.cn

UK

• Email: technical@abcam.com

• Tel: +44 (0) 1223 696000

• Website: www.abcam.com

Japan

• Email: technical@abcam.co.jp

• 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

• sales@abcam.com

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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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