Description

pgRNA_genes.sh is a wrapper for scripts that are used to:

• browse 2kb upstream and downstream regions of the targets mentioned in the input bed file
• identify guide sequences with PAM sites
• mapping the guides across the genome to identify potential off target
• match pairs of guide from upstream and downstream the target to filter them on the basis of GC content
• parse the alignment map to filter and sort on the basis of PAM, mismatches and GC content
• Subset top 4 pairs of non-redundant guide RNA pairs

Pre-requisites

• Install bowtie, samtools
• Create a src directory

    mkdir src
    cd src
    wget ftp://ftp.ncbi.nlm.nih.gov/genomes/all/GCF/003/668/045/GCF_003668045.1_CriGri-PICR/GCF_003668045.1_CriGri-PICR_genomic.fna.gz
    gunzip GCF_003668045.1_CriGri-PICR_genomic.fna.gz 
    mv GCF_003668045.1_CriGri-PICR_genomic.fna picr.fa

• Create fasta index with samtools for the reference sequence

    samtools faidx picr.fa

• Create bowtie index for reference sequence

    bowtie-build picr.fa picr

Usage

./pgRNA_genes.sh -bed <input bed file>

Input bed file: Its a tab separated text file with ".bed" extension

chr	start	stop	name	score	strand