Analysis software for multi-target CRISPR

Companion code for:

Zou, R.S., Marin-Gonzalez, A., Liu, Y., Liu, H.B., Shen, L., Dveirin, R., Luo, J.X., Kalhor, R. and Ha, T. Massively parallel genomic perturbations with multi-target CRISPR reveal new insights on Cas9 activity and DNA damage responses at endogenous sites. bioRxiv (2022). https://www.biorxiv.org/content/10.1101/2022.01.18.476836v1

Software requirements

• Anaconda Python 3.7 (Anaconda's python distribution comes with the required numpy and scipy libraries)
• pysam
• bowtie2
• samtools
• Ensure that both samtools and bowtie2 are added to path and can be called directly from bash

Data requirements

• https://www.ncbi.nlm.nih.gov/bioproject/PRJNA733683

Installation

1. Download sequencing reads in FASTQ format from SRA
2. Download the prebuilt bowtie2 indices for human hg19 and hg38 genome assemblies
   • Human hg38
   • Human hg19
   • Extract from archive, move to the corresponding folders named hg38_bowtie2/ and hg19_bowtie2/
3. Download two human hg19 and hg38 genome assemblies in FASTA format
   • hg38.fa
   • hg19.fa
   • Extract from archive, move to the corresponding folders named hg38_bowtie2/ and hg19_bowtie2/
4. Generate FASTA file indices
   • samtools faidx hg38_bowtie2/hg38.fa
   • samtools faidx hg19_bowtie2/hg19.fa

Usage

Bash scripts are used to automate the processing of sequencing data.

Python scripts are used to perform analysis of various data featured in the manuscript. They are labeled script_*_*.py, such as script_1_putative.py

List of mgRNA sequences

List of target sites for the 'GG', 'CT', and 'TA' mgRNA sequences, along with its hg38 genomic context, are included in mgRNA_target_sites.xlsx.