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DpnII-seq

snakemake-run

DpnII-seq is an experimental assay to measure the genome wide cutting frequency of the restriction enzyme DpnII. Originally DpnII-seq was developed alongside Liquid Chromatin Hi-C to control for biases in cutting frequency when estimating chromatin contact stability in K562 cells. However, the DpnII-seq assay and analysis pipeline could be modified for analyzing cutting frequency for variable restriction enzymes in variable cell types. The DpnII-seq workflow performs mapping, filtering of artifacts, multiple resolution binning, copy number correction and plotting of quality metrics.

This workflow has options for digesting with the following restriction enzymes:

As K562 cells have a primarily triploid karyotype with regions of variable copy number, the analysis workflow corrects coverage tracks to a diploid state genome wide. If the user is applying DpnII-seq to cells with variable copy number states we provide scripts to correct for this bias using a Gaussian mixture model approach.

Experimental Protocol

dpn

Computational Workflow

dag

Requirements

Install Snakemake via Miniconda here

Set up channels
conda config --add channels defaults
conda config --add channels conda-forge
conda config --add channels bioconda
conda config --add channels r
Set up environment
conda env create --file envs/DpnII-seq_env.yaml
source activate DpnII-seq
Set up data directory
.
├── data
│   ├── binning
│   ├── copy_number
│   ├── dpnII_sites
│   ├── fastq
│   ├── fatI_sites
│   ├── hindIII_sites
│   └── indexes
├── DpnII-seq

DpnII-seq workflow requires a data directory with the following files:

  • binning/
    • bedfiles denoting binned genome (ex. hg19_40kb.bed)
  • copy_number/
    • bedfiles denoting binned genome with copy number state (4th column)
  • [X]_sites/
    • bedfiles denoting restriction sites of restriction enzyme X for each chromosome (ex. dpnII_sites_chr1.bed)
  • fastq/
    • paired end fastq files (*_R1.fastq, *_R2.fastq)
  • indexes/
    • bowtie-build index files for reference genome (*.ebwt)

Usage

Currently the DpnII-seq workflow has only been tested in an LSF cluster environment

Snake Command
snakemake -j 10 --latency-wait 60 --cluster-config cluster.json --cluster "bsub -q {cluster.queue} -W {cluster.time} -R {cluster.memory} -n {cluster.cores} -o {cluster.output} -e {cluster.error}" -p

Contributing to DpnII-seq

All contributions, bug reports, bug fixes, documentation improvements, enhancements, and ideas are welcome!

References

Houda Belaghzal*, Tyler Borrman*, Andrew D. Stephens, Denis L. Lafontaine, Sergey V. Venev, Zhiping Weng, John F. Marko, Job Dekker. (2019). Compartment-dependent chromatin interaction dynamics revealed by liquid chromatin Hi-C. bioRxiv

License

GNU General Public License v3.0