BashRegion

This function takes as input an Interval-format file and output all sgRNA sites in the interval in both CSV and FASTA format.

Author: Neville Sanjana ([email protected])

CHANGELOG:

• December 2015

  • Support for 5' PAMs (e.g. Cpf1)

• June 2015

  • Consolidated modules
  • Prepared for public release

• November 2014

  • Initial version

USAGE REQUIREMENTS

Python 2.7 with the following libraries installed:

• Default Python libraries (sys, os, csv, urllib2, math, ElementTree)
• BioConductor (for SeqIO, SeqUtils, Seq, IUPAC)
• numpy
• pandas

Please make sure all of these libraries are installed and working before proceeding.

To run, place the BashRegion.py in a directory that also includes the input file (specified in intervalFile_INPUT). Then run the following command

python BashRegion.py

INPUT/PARAMETERS TO SET:

1. intervalFile_INPUT : A tab-delimited file (such as the included "SampleInput.interval" file) with a list of coordinates. See comment in the PARAMETERS section for an example of the format.

2. referenceGenomeForDAS : The reference genome to use for translating the genomic coordinates in intervalFile_INPUT to genomic DNA sequence. For the included file ("SampleInput.interval"), make sure to set referenceGenomeForDAS = "hg19" since the coordinates are from the hg19 human reference genome.

3. outDirectory : The name of the directory for output files. It will be created if not already present.

4. outShortName : A short string to prefix the output files with.

5. cutoffSpacing : The minimum spacing between selected sgRNAs on the same strand. For example, if cutoffSpacing = 2, then the next sgRNA must be > 2 bp from the previous sgRNA chosen. To find all sgRNAs in a region, set cutoffSpacing = 0.

6. PAM : A string that contains the PAM sequence (e.g. "NGG" for SpCas9)

FORMAT OF intervalFile_INPUT:

The interval file should have 5 columns and be tab-delimited

<chromosome> <strand (optional but needs column to be there)> <base start> <base end> <geneID>

Example of intervalFile_INPUT:

chr12 . 51057788 51173928 ATF1

chr1 + 8821058 8921418 ENO1

Note that the strand parameter is not used by BashRegion.py but placed here for compatibility with standard interval file formats.

OUTPUT FILES:

Output is all sgRNAs on both strands in CSV and FASTA format.

The CSV format output includes the sgRNA sequences and the following parameters:

• Spacer number
• Strand
• Start Pos
• End Pos
• Chromosome
• ChrStart
• ChrEnd
• Cut Site
• Distance from Previous Spacer
• Spacer (i.e. sgRNA)
• PAM
• Spacer GC
• Seq ID