HiC-Pro: An optimized and flexible pipeline for Hi-C data processing
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README.rst

HiC-Pro Quick Start Guide

This page is just a quick start guide, please read the full online manual for more information !

See NEWS for information about changes in this and previous versions

See LOGBOOK for details about the HiC-Pro developments

See LICENSE for license information

Where to get help ?

For any question about HiC-Pro, please contact nicolas.servant@curie.fr or use the HiC-Pro forum !

What is HiC-Pro ?

HiC-Pro was designed to process Hi-C data, from raw fastq files (paired-end Illumina data) to the normalized contact maps. Since version 2.7.0, HiC-Pro supports the main Hi-C protocols, including digestion protocols as well as protocols that do not require restriction enzyme such as DNase Hi-C. In practice, HiC-Pro can be used to process dilution Hi-C, in situ Hi-C, DNase Hi-C, Micro-C, capture-C, capture Hi-C or HiChip data.
The pipeline is flexible, scalable and optimized. It can operate either on a single laptop or on a computational cluster. HiC-Pro is sequential and each step of the workflow can be run independantly.
HiC-Pro includes a fast implementatation of the iterative correction method (see the iced python library for more information).
In addition, HiC-Pro can use phasing data to build :ref:`allele specific contact maps <AS>`.

If you use HiC-Pro, please cite :

HiC-Pro: An optimized and flexible pipeline for Hi-C processing. Servant N., Varoquaux N., Lajoie BR., Viara E., Chen CJ., Vert JP., Dekker J., Heard E., Barillot E. Genome Biology 2015, 16:259 doi:10.1186/s13059-015-0831-x http://www.genomebiology.com/2015/16/1/259

How to install it ?

The HiC-Pro pipeline requires the following dependencies :

  1. The bowtie2 mapper
  2. Python (>2.7, python-3 is not supported) with pysam (>=0.8.3), bx-python(>=0.5.0), numpy(>=1.8.2), and scipy(>=0.15.1) libraries
  3. R with the RColorBrewer and ggplot2 (>2.2.1) packages
  4. g++ compiler
  5. samtools (>1.1)
  6. Unix sort (which support -V option) is required ! For Mac OS user, please install the GNU core utilities !

Bowtie >2.2.2 is strongly recommanded for allele specific analysis.

To install HiC-Pro (>=2.7.8), be sure to have the appropriate rights and run :

tar -zxvf HiC-Pro-master.tar.gz
cd HiC-Pro-master
## Edit config-install.txt file if necessary
make configure
make install
For older version (<2.7.8), the following process can be used
tar -zxvf HiC-Pro-master.tar.gz
cd HiC-Pro-master
## Edit config-install.txt file if necessary
make CONFIG_SYS=config-install.txt install
Note that if some of these dependencies are not installed (i.e. not detected in the $PATH), HiC-Pro will try to install them.
You can also edit the config-install.txt file and manually defined the paths to dependencies.
SYSTEM CONFIGURATION
PREFIX Path to installation folder
BOWTIE2_PATH Full path the bowtie2 installation directory
SAMTOOLS_PATH Full path to the samtools installation directory (>1.1 )
R_PATH Full path to the R installation directory
PYTHON_PATH Full path to the python installation directory (>2.7 - python3 not supported)
CLUSTER_SYS Scheduler to use for cluster submission. Must be TORQUE, SGE, SLURM or LSF

Annotation Files

In order to process the raw data, HiC-Pro requires three annotation files. Note that the pipeline is provided with some Human and Mouse annotation files. Please be sure that the chromosome names are the same than the ones used in your bowtie indexes !

  1. A BED file of the restriction fragments after digestion. This file depends both of the restriction enzyme and the reference genome. See the :ref:`FAQ <FAQ>` and the :ref:`HiC-Pro utilities <UTILS>` for details about how to generate this file. A few annotation files are provided with the HiC-Pro sources as examples.
chr1   0       16007   HIC_chr1_1    0   +
chr1   16007   24571   HIC_chr1_2    0   +
chr1   24571   27981   HIC_chr1_3    0   +
chr1   27981   30429   HIC_chr1_4    0   +
chr1   30429   32153   HIC_chr1_5    0   +
chr1   32153   32774   HIC_chr1_6    0   +
chr1   32774   37752   HIC_chr1_7    0   +
chr1   37752   38369   HIC_chr1_8    0   +
chr1   38369   38791   HIC_chr1_9    0   +
chr1   38791   39255   HIC_chr1_10   0   +
(...)
  1. A table file of chromosomes' size. This file can be easily find on the UCSC genome browser. Of note, pay attention to the contigs or scaffolds, and be aware that HiC-pro will generate a map per chromosomes pair. For model organisms such as Human or Mouse, which are well annotated, we usually recommand to remove all scaffolds.
chr1    249250621
chr2    243199373
chr3    198022430
chr4    191154276
chr5    180915260
chr6    171115067
chr7    159138663
chr8    146364022
chr9    141213431
chr10   135534747
(...)
  1. The bowtie2 indexes. See the bowtie2 manual page for details about how to create such indexes.

How to use it ?

  1. First have a look at the help message !
HiC-Pro --help
usage : HiC-Pro -i INPUT -o OUTPUT -c CONFIG [-s ANALYSIS_STEP] [-p] [-h] [-v]
Use option -h|--help for more information

HiC-Pro 2.10.0
---------------
OPTIONS

 -i|--input INPUT : input data folder; Must contains a folder per sample with input files
 -o|--output OUTPUT : output folder
 -c|--conf CONFIG : configuration file for Hi-C processing
 [-p|--parallel] : if specified run HiC-Pro on a cluster
 [-s|--step ANALYSIS_STEP] : run only a subset of the HiC-Pro workflow; if not specified the complete workflow is run
    mapping: perform reads alignment
    proc_hic: perform Hi-C filtering
    quality_checks: run Hi-C quality control plots
    build_contact_maps: build raw inter/intrachromosomal contact maps
    ice_norm: run ICE normalization on contact maps
 [-h|--help]: help
 [-v|--version]: version
  1. Copy and edit the configuration file 'config-hicpro.txt' in your local folder. See the :ref:`manual <MANUAL>` for details about the configuration file
  2. Put all input files in a rawdata folder. The input files have to be organized with one folder per sample, with ;
+ PATH_TO_MY_DATA
  + sample1
    ++ file1_R1.fastq.gz
    ++ file1_R2.fastq.gz
    ++ ...
  + sample2
    ++ file1_R1.fastq.gz
    ++ file1_R2.fastq.gz
  *...
  1. Run HiC-Pro
  • On your laptop
MY_INSTALL_PATH/bin/HiC-Pro -i FULL_PATH_TO_DATA_FOLDER -o FULL_PATH_TO_OUTPUTS -c MY_LOCAL_CONFIG_FILE
  • Using a cluster (TORQUE/SGE/SLURM/LSF)
MY_INSTALL_PATH/bin/HiC-Pro -i FULL_PATH_TO_DATA_FOLDER -o FULL_PATH_TO_OUTPUTS -c MY_LOCAL_CONFIG_FILE -p

You will get the following message :

Please run HiC-Pro in two steps :
1- The following command will launch the parallel workflow through 12 torque jobs:
qsub HiCPro_step1.sh
2- The second command will merge all outputs to generate the contact maps:
qsub HiCPro_step2.sh

Execute the displayed command from the output folder:

qsub HiCPro_step1.sh
774410[].torque.curie.fr

Then wait for the torque mails... :) Once executed succesfully (may take several hours), then type:

qsub HiCPro_step2.sh

Test Dataset

The test dataset and associated results are available at https://zerkalo.curie.fr/partage/HiC-Pro/. Small fastq files (2M reads) extracted from the Dixon et al. 2012 paper are available for test.

## Get the data. Will download a test_data folder and a configuration file
wget https://zerkalo.curie.fr/partage/HiC-Pro/HiCPro_testdata.tar.gz && tar -zxvf HiCPro_testdata.tar.gz

## Edit the configuration file and set the path to Human bowtie2 indexes

## Run HiC-Pro

time HICPRO_INSTALL_DIR/bin/HiC-Pro -c config_test_latest.txt -i test_data -o hicpro_latest_test

Run HiC-Pro 2.10.0
--------------------------------------------
vendredi 22 décembre 2017, 13:34:18 (UTC+0100)
Bowtie2 alignment step1 ...
/home/nservant/Apps/HiC-Pro_2.10.0/scripts/bowtie_wrap.sh -c /home/nservant/Desktop/hicpro_dev/test-op/config_test_latest.txt -u >> hicpro.log
--------------------------------------------
vendredi 22 décembre 2017, 13:34:55 (UTC+0100)
Bowtie2 alignment step2 ...
/home/nservant/Apps/HiC-Pro_2.10.0/scripts/bowtie_wrap.sh -c /home/nservant/Desktop/hicpro_dev/test-op/config_test_latest.txt -l >> hicpro.log
--------------------------------------------
vendredi 22 décembre 2017, 13:35:05 (UTC+0100)
Combine both alignment ...
/home/nservant/Apps/HiC-Pro_2.10.0/scripts/bowtie_combine.sh -c /home/nservant/Desktop/hicpro_dev/test-op/config_test_latest.txt >> hicpro.log
--------------------------------------------
vendredi 22 décembre 2017, 13:35:09 (UTC+0100)
Bowtie2 mapping statistics for R1 and R2 tags ...
/home/nservant/Apps/HiC-Pro_2.10.0/scripts/mapping_stat.sh -c /home/nservant/Desktop/hicpro_dev/test-op/config_test_latest.txt >> hicpro.log
--------------------------------------------
vendredi 22 décembre 2017, 13:35:10 (UTC+0100)
Pairing of R1 and R2 tags ...
/home/nservant/Apps/HiC-Pro_2.10.0/scripts/bowtie_pairing.sh -c /home/nservant/Desktop/hicpro_dev/test-op/config_test_latest.txt >> hicpro.log
--------------------------------------------
vendredi 22 décembre 2017, 13:35:16 (UTC+0100)
Assign alignments to restriction fragments ...
/home/nservant/Apps/HiC-Pro_2.10.0/scripts/mapped_2hic_fragments.sh -c /home/nservant/Desktop/hicpro_dev/test-op/config_test_latest.txt >> hicpro.log
--------------------------------------------
vendredi 22 décembre 2017, 13:35:45 (UTC+0100)
Merge multiple files from the same sample ...
/home/nservant/Apps/HiC-Pro_2.10.0/scripts/merge_valid_interactions.sh -c /home/nservant/Desktop/hicpro_dev/test-op/config_test_latest.txt >> hicpro.log
--------------------------------------------
vendredi 22 décembre 2017, 13:35:46 (UTC+0100)
Merge stat files per sample ...
/home/nservant/Apps/HiC-Pro_2.10.0/scripts/merge_stats.sh -c /home/nservant/Desktop/hicpro_dev/test-op/config_test_latest.txt >> hicpro.log
--------------------------------------------
vendredi 22 décembre 2017, 13:35:46 (UTC+0100)
Run quality checks for all samples ...
/home/nservant/Apps/HiC-Pro_2.10.0/scripts/make_plots.sh -c /home/nservant/Desktop/hicpro_dev/test-op/config_test_latest.txt -p "all" >> hicpro.log
--------------------------------------------
vendredi 22 décembre 2017, 13:35:55 (UTC+0100)
Generate binned matrix files ...
/home/nservant/Apps/HiC-Pro_2.10.0/scripts/build_raw_maps.sh -c /home/nservant/Desktop/hicpro_dev/test-op/config_test_latest.txt
--------------------------------------------
vendredi 22 décembre 2017, 13:35:56 (UTC+0100)
Run ICE Normalization ...
/home/nservant/Apps/HiC-Pro_2.10.0/scripts/ice_norm.sh -c /home/nservant/Desktop/hicpro_dev/test-op/config_test_latest.txt >> hicpro.log

real1m38.855s
user3m13.344s
sys0m31.432s

Using HiC-Pro in a Singularity container

HiC-Pro v2.10.0 provides a Singularity container to ease its installation process. A ready-to-use container is available here.

1- Install singularity

2- Build the singularity HiC-Pro image using the 'Singularity' file available in the HiC-Pro root directory.

singularity create -s 5000 hicpro_latest_ubuntu.img
sudo singularity -d bootstrap hicpro_latest_ubuntu.img MY_INSTALL_PATH/HiC-Pro/Singularity

3- Run HiC-pro

singularity exec hicpro_latest_ubuntu.img HiC-Pro -h