sgRNA design for disrupting TFBS

usage: gRNA_design_TF.py [-h] [-j JID] -TAD TAD_FILE -peak PEAK_FILE -motif
                         MOTIF_FILE -gene GENE_FILE -off_target NUM_MATCH_FILE
                         [-l FLANKING_LENGTH] [--PAM_seq PAM_SEQ]
                         [--motif_position_anchor MOTIF_POSITION_ANCHOR]
                         [-g GENOME] [--genome_fasta GENOME_FASTA]

optional arguments:
  -h, --help            show this help message and exit
  -j JID, --jid JID     enter a job ID, which is used to make a new directory.
                        Every output will be moved into this folder. (default:
                        gRNA_design_TF_yli11_2020-12-06)
  -l FLANKING_LENGTH, --flanking_length FLANKING_LENGTH
                        number of bp flanking the motif bed file (default: 25)
  --PAM_seq PAM_SEQ     PAM seq (default: NGG)
  --motif_position_anchor MOTIF_POSITION_ANCHOR
                        which position to use as +1 position, default is for
                        WGATAR, where the first A is used as +1 then T is -1
                        (default: 3)

required named arguments:
  -TAD TAD_FILE, --TAD_file TAD_FILE
                        bed file, at least 4 columns (default: None)
  -peak PEAK_FILE, --peak_file PEAK_FILE
                        bed file, at least 4 columns (default: None)
  -motif MOTIF_FILE, --motif_file MOTIF_FILE
                        bed file, at 6 columns, containing strand (default:
                        None)
  -gene GENE_FILE, --gene_file GENE_FILE
                        bed file, at least 4 columns (default: None)
  -off_target NUM_MATCH_FILE, --Num_match_file NUM_MATCH_FILE
                        tsv file, 2 columns, sgRNA seq and number of matches
                        in the genome (default: None)

Genome Info:
  -g GENOME, --genome GENOME
                        genome version: hg19, hg38, mm9, mm10. By default,
                        specifying a genome version will automatically update
                        chrom size. (default: hg19)
  --genome_fasta GENOME_FASTA
                        genome version: hs, mm (default:
                        /home/yli11/Data/Human/hg19/fasta/hg19.fa)

Summary

This pipeline was developed in the GATA1 disruption project, but is generic for other sgRNA design projects focusing on TF motifs.

Input

You will need 4 bed files and 1 tsv file:

1. bed files for TF peaks, TF motifs, TADs, and gene bed file. Note that TF motif bed file requires 6 columns, where the last column is strand.

2. a tsv file for sgRNA genome-wide number of occurrences, first column is sgRNA_seq, second column is an integer.

Note

TAD.bed, gene.bed, peak.bed requires 4 columns, the last column is id or name. motif.bed requires 6 columns: chr, start, end, name, value, strand.

Usage

hpcf_interactive

module load python/2.7.13

gRNA_design_TF.py -TAD tad.bed -peak GATA1_peaks.bed -motif WGATAR.bed -gene ranked_genes.bed -off_target number_matches.bed -g hg19 --PAM NG -l 25 --motif_position_anchor 3

Output

The final output file is: Motif_{flanking_length}_gRNA.annot.bed.rel_edit_pos.filter.bed

It was filtered based on:

• 1. only 1 match in the genome

• 2. GC% : 0.2 - 0.8

• 3. at least one editable base in the editing window (predefined) 3-8

The output columns are:

0-5: sgRNA bed file
6-11: motif bed file
12-15: peak bed file
16: TAD name
17: gene name
18: relative position to the anchor ``--motif_position_anchor``
19: number of editable bases
20: number of off-targets, -1 means not found in the provided off-target table

Comments

code @ github.