ISMB Tutorial: 3D Genome Data Processing, Analysis, and Visualization Tutorial

Files in this repository

• Tutorial Part 1 (Introduction to HiC): ISMB-Tutorial-Part-1-Nezar.pptx
• Tutorial Part 2 (Data Processing and Visualization): Introduction and ISMB-Tutorial-Part-2-Soo-Nils-Peter.html or https://hms-dbmi.github.io/3d-genome-processing-tutorial/
• Tutorial Part 3 (Data Analysis for Nuclear Compartmentalization): ISMB-Tutorial-Part-3-Ma.pdf

Organizers & Presenters

• Nezar Abdennur, PhD student, MIT
• Soo Lee, Senior Bioinformatics Scientist, Harvard Medical School
• Nils Gehlenborg, Assistant Professor, Harvard Medical School
• Peter Kerpedjiev, Postdoctoral Research Fellow, Harvard Medical School
• Jian Ma, Associate Professor, Carnegie Mellon University

Motivation

Due in large part to the explanatory power of chromosome organization in gene regulation, its association with disease and disorder as well as the unanswered questions regarding the mechanisms behind its maintenance and function, the 3D structure and function of the genome are becoming increasingly target of scientific scrutiny. With efforts such as the 4D Nucleome Project and ENCODE 4 already beginning to generate large amounts of data, the ability to analyze and visualize it will be a valuable asset to any computational biologist tasked with interpretation of experimental results.

Objectives

The objectives of this tutorial are

• To introduce the theoretical concepts related to 3D genome data analysis
• To familiarize participants with the data types, analysis pipeline, and common tools for analysis and visualization of 3D genome data
• To provide a hands on experience in data analysis by walking through some common use cases of existing tools for data analysis and visualization.

Goal

After the workshop participants should be able to obtain, process, analyze, and visualize 3D genome data on their own as well as to understand some of the logic, motivation and pitfalls associated with common operations such as matrix balancing and multi-resolution visualization.

Intended audience and level

The subject matter and practical exercises presented in this tutorial will be accessible to a broad audience. Prior experience with next generation sequencing and the data it produces will be helpful for understanding the subsequent processing steps used to derive contact maps as well as some of the artifacts that can arise during data processing.

The material will be most useful to computational biologists and biologists working on genomics-related topics. Tentative Schedu

Student Requirements:

• Install Docker
  • https://www.docker.com/community-edition
• Install Miniconda
  • https://conda.io/miniconda.html
• Windows users
  • Putty (for ssh)
  • https://www.chiark.greenend.org.uk/~sgtatham/putty/latest.html

Agenda

10:00 - 10:25 - Introduction and Overview

• Who are we? [5 minutes]
• What we’ll cover [5 minutes]
• AWS accounts and ssh in [10 minutes]

10:25 - 11:30 Hi-C Analysis

      Intro to Hi-C [Nezar]

• 15 minutes:

  • Introduction
    • Protocol
    • What Hi-C sees
    • Levels of genome organization inferred from patterns
      • Checkering: compartments
      • Enriched squares without checkering: TADs
      • Corner peaks: loops
    • How Hi-C is processed
    • How features are assessed
  • Hi-C processing steps (informational)
    • Mapping
    • Filtering
    • Creating a list of contacts
    • Binning
    • Normalization
    • Feature analysis
      • scaling
      • compartments
      • TADs
    • QC

• 45min

  • Practical - processing pipeline [Soo]:
    • Mapping
      • bwa mem -SP5M index fastq1 fastq2 | samtools view -bhS - > output.bam
      • samtools view output.bam | head -n 5
      • output: bam file
    • Filtering / sorting / Creating a list of contacts
      • pairsamtools
      • outputs: pairs, bam
      • Pairs / pairix (indexes the pairs file)
    • Binning
      • Cooler / cool
    • Normalization
      • Cooler / cool
    • map a small Hi-C dataset using distiller (https://github.com/mirnylab/distiller) and generate contact matrices using cooler (https://github.com/mirnylab/cooler)
  • Practical - Feature analysis [Nezar]:
    • Jupyter notebook cooler walkthrough
    • Cis vs trans and scaling (contact probability vs genomic distance)
    • Compartment profile, saddle plots
    • Insulation, TADs
    • Pileups

      11:30 - 11:45 Coffee Break

      11:45 - 12:55 Visualization

      Existing tools for contact matrix exploration

• 20 minutes [Nils]:
  • 3D genome browser
  • WashU epigenome browser
  • Juicebox
  • HiGlass

      Using HiGlass (http://higlass.io) to display contact maps [Peter]

• 30 minutes:
  • Overview of common operations such as adding tracks, removing tracks, adding views, removing view, linking views by zoom and location
  • Practical:
    • Create an interactive version of a figure
• 20 minutes: Installing HiGlass
  • Overview of the HiGlass architecture and description of the infrastructure used to run it
  • Practical:
    • Create a local HiGlass instance
    • Convert a contact map to multi-resolution format and import it
    • Convert a bigWig file to hitile format and import it
    • Open both files in the client and navigate to an interesting location

      12:55 - 1:30 - Data Analysis for Nuclear Compartmentalization [Jian Ma]

• Introduction
• DamID analysis
• Repli-seq analysis
• Data from emerging technologies

Instructor Bios

Nezar Abdennur

Nezar is a PhD candidate in Computational and Systems Biology at MIT. His research focuses on the determinants of 3D genome organization and the development of tools for dealing with large Hi-C datasets. http://nvictus.me

Soo Lee

Soo Lee is a Senior Bioinformatics Scientist in the Department of Biomedical Informatics at Harvard Medical School. She is creating cloud-based pipelines for Hi-C and other genomic data and developing infrastructure for automation of such pipelines as part of the 4D Nucleome Data Coordination and Integration Center. https://compbio.hms.harvard.edu/people/soohyun-lee

Nils Gehlenborg

Nils is an Assistant Professor in the Department of Biomedical Informatics at Harvard Medical School. His research group is developing tools to visualize 3D genome conformation data as well as heterogeneous data from large-scale cancer genomics studies. http://gehlenborglab.org

Peter Kerpedjiev

Peter is a postdoctoral researcher working on creating tools (such as HiGlass) for visualizing large genomic data sets. Web site: http://emptypipes.org/about

Jian Ma

Jian Ma is an Associate Professor in the School of Computer Science at Carnegie Mellon University. The research in his group focuses on developing algorithms to better understand genome structure and function. http://www.cs.cmu.edu/~jianma/

Resources

Software

• bwa and SAM spec
• pairsamtools
• pairix
• cooler and docs
• HiGlass and wiki
• HiPiler

Bioinformatics workflow managers

• snakemake
• nextflow

Package and environment management

• conda
• bioconda

Hi-C methods papers

• Imakaev, Maxim, et al. "Iterative correction of Hi-C data reveals hallmarks of chromosome organization." Nature methods 9.10 (2012): 999-1003. doi:10.1038/nmeth.2148
• Lajoie, Bryan R., Job Dekker, and Noam Kaplan. "The Hitchhiker’s guide to Hi-C analysis: practical guidelines." Methods 72 (2015): 65-75. doi:10.1016/j.ymeth.2014.10.031