TAD-and-loop-identification-workflows

The repository contains the code to identify TADs and loops from Hi-C and HiChIP library

The flowchart

The complete process of identifying TADs and loops by multiple software

By running "sh workflow.sh", users can directly identify TADs through TADLib and Juicer, identify loops through Fit-Hi-C, Juicer and FitHiChIP.

The test data of TADLib is ./input/TADLib_input_data/chr5_chr6.cool.gz. The test data need to decompress before runing
The test data of Juicer is ./input/juicer_input_data/chr1.hic.gz. The test data need to decompress before runing
The test data of Fit-Hi-C is in ./input/fithic_input_data
The test data of FitHiChIP is ./input/FitHiChIP_input_data/chr21.allValidPairs.gz. The test data need to decompress before running

Limited by file size of github, the rawdata of HiC-Pro and hichipper need to be downloaded by user

The running methods of HiC-Pro and hichipper are shown in workflow.sh
The downloading detail of HiC-Pro rawdata is shown in lib/HiC-Pro/Rawdata/README
The downloading detail of hichipper rawdata is shown in lib/hichipper/HiC-Pro/Rawdata/README

sh workflow.sh

Rawdata

The input/rawdata.txt includes the download adderss of rawdata used in this pipeline

Detailed steps of HiC-Pro, Juicer, TADLib, Fit-Hi-C, FitHiChIP, hichipper

Lib folder contains some scripts required to run this pipeline

HiC-Pro needs downloaded rawdata, and the detail can be found in ./lib/HiC-Pro/run_HiC-Pro.sh and ./lib/HiC-Pro/Rawdata/README

## Enzyme fragments
python ./annotation/digest_genome.py ../Rawdata/Gbar.fa -r dpnii -o ./annotation/Gbar_dpnii.bed

## Chromosome size
cd annotation
samtools faidx Gbar.fa
awk -v OFS="\t" '{print $1,$2}' Gbar.fa.fai > ./annotation/chrom_Gb.sizes
cd ..

## Bowtie2
cd Bowtie2
ln -s ../annotation/Gbar.fa
bowtie2-build -f Gbar.fa Gbar
cd ..


## Personal computer
HiC-Pro -c HiC-Pro.config -i ../Rawdata -o result 

## High performance computing
HiC-Pro -c HiC-Pro.config -i ../Rawdata -o result -p
cd result
sh HiCPro_step1.sh
sh HiCPro_step2.sh

More parameter details of HiC-Pro can be found in https://nservant.github.io/HiC-Pro/

Juicer, identify TADs and infer loops

## Identify TADs
java -jar scripts/juicer_tools.jar arrowhead -m 2000 -r 20000 -k KR --threads 15 ../../input/juicer_input_data/chr1.hic ../../output/Juicer/TAD

## Infer loops
java -jar scripts/juicer_tools.jar hiccups --cpu -r 5000,10000 -f 0.1,0.1 -p 4,2 -i 7,5 -d 20000,20000 -t 0.02,1.5,1.75,2 -k KR --threads 10 ../../input/juicer_input_data/chr1.hic ../../output/Juicer/loop

More parameter details of Juicer can be found in https://github.com/aidenlab/juicer/wiki

TADLib

## Produce cool file
cp ../../input/HiC-Pro_result_matrix_chr5_chr6/20Kb/chr5.matrix ../../input/TADLib_input_data/1_1.txt
cp ../../input/HiC-Pro_result_matrix_chr5_chr6/20Kb/chr6.matrix ../../input/TADLib_input_data/2_2.txt

## toCooler from HiCPeaks
toCooler -O ../../input/TADLib_input_data/chr5_chr6.cool -d ./chr5_chr6_dataset --chromsizes-file ./chr5_chr6.chromsizes --nproc 2 --no-balance

## Identify TADs
hitad -O ../../output/TAD_Lib/cotton_chr5_chr6_TADLib_TAD.bed -d meta_file --logFile hitad.log -p 2 -W RAW 

More parameter details of TADLib can be found in https://xiaotaowang.github.io/TADLib/hitad_api.html

Fit-Hi-C

## Convert the result file produce by HiC-Pro to the input file of Fit-Hi-C
python hicpro2fithic.py -i ../../input/HiC-Pro_result_matrix_chr5_chr6/5Kb/chr5.matrix -b ../../input/HiC-Pro_result_matrix_chr5_chr6/5Kb/chr5_abs.bed -r 5000 -o ../../input/fithic_input_data -n chr5 
python hicpro2fithic.py -i ../../input/HiC-Pro_result_matrix_chr5_chr6/5Kb/chr6.matrix -b ../../input/HiC-Pro_result_matrix_chr5_chr6/5Kb/chr6_abs.bed -r 5000 -o ../../input/fithic_input_data -n chr6 

## Infer loops
fithic -f ../../input/fithic_input/input_data/chr5.fithic.fragmentMappability.gz -i ../../input/fithic_input_data/chr5.fithic.interactionCounts.gz -r 5000 -L 10000 -U 3000000 -p 2 -b 200 -o ../../output/Fit-Hi-C -l chr5
fithic -f ../../input/fithic_input/input_data/chr6.fithic.fragmentMappability.gz -i ../../input/fithic_input_data/chr6.fithic.interactionCounts.gz -r 5000 -L 10000 -U 3000000 -p 2 -b 200 -o ../../output/Fit-Hi-C -l chr6

More parameter details of Fit-Hi-C can be found in https://github.com/ay-lab/fithic

FitHiChIP

## Infer loops
bash FitHiChIP_HiCPro.sh -C configfile_BiasCorrection_CoverageBias_chr21 

More parameter details of FitHiChIP can be found in https://ay-lab.github.io/FitHiChIP/

hichipper needs output file produced by HiC-Pro

HiC-Pro needs downloaded rawdata. The details can be found in hichipper/HiC-Pro/annotation/README and hichipper/HiC-Pro/Rawdata/README

## run HiC-Pro

## Enzyme fragments
python ./annotation/digest_genome.py ../Rawdata/GRCh38.fa -r dpnii -o ./annotation/GRCh38_dpnii.bed

## Chromosome size
cd annotation
samtools faidx GRCh38.fa
awk -v OFS="\t" '{print $1,$2}' GRCh38.fa.fai > GRCh38_sizes.txt
cd ..

## Bowtie2
cd Bowtie2
ln -s ../annotation/GRCh38.fa
bowtie2-build -f GRCh38.fa GRCh38
cd ..

## Personal computer
HiC-Pro -c HiC-Pro.config -i ../Rawdata -o result 

## High performance computing
HiC-Pro -c HiC-Pro.config -i ../Rawdata -o result -p
cd result
sh HiCPro_step1.sh
sh HiCPro_step2.sh

## running hichipper
hichipper --out result two.yaml --macs2-string "-q 0.1" --macs2-genome "hs"

More parameter details of hichipper can be found in https://hichipper.readthedocs.io/en/latest/

Output

The folder contains the results of TADs and loops from Hi-C and HiChIP library

Result visualisation

workflow.sh

The script records the commands for running theses tools