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guide_design_tools

Tools for automated design of CRISPR guides for DASH, FLASH and CRISPR tagging experiments.

DASHit

DASHit is a software package to design guide RNAs for DASH experiments.

Installing

DASHit runs on macOS and Linux (it’s likely DASHit will compile on Windows, but we haven’t tested this!). To install DASHit, you’ll need:

Dependencies

  1. [go|https://www.golang.org] Note: brew install golang, nix-env -i go
  2. [Python 3.7|https://www.python.org/] Note: brew install

Installing DASHit

  1. Install the Biohub’s [special_ops_crispr_tools|https://www.github.com/czbiohub/special_ops_crispr_tools/]
    git clone https://github.com/czbiohub/special_ops_crispr_tools.git
    cd special_ops_cripr_tools
    make
    sudo make install
    

    Note: Use, e.g., PREFIX=$HOME make install to install special_ops_crispr_tools in a different location: you’ll just need the crispr_sites and offtarget binaries in your path

  2. Create and enter a Python virutal environment:
    python -m venv ~/.virtualenvs/dashit
    source ~/.virtualenvs/dashit/bin/activate
    
  3. Install DASHit
    git clone https://github.com/czbiohub/guide_design_tools
    cd guide_design_tools
    make install
    

Running DASHit

Once DASHit is installed, run it from the DASHit virtual environment.

source ~/.virtualenvs/dashit/bin/activate
dashit-reads-filter --help
crispr_sites -h
offtarget -h
optimize_guides -h

Designing guides

  1. Run crispr_sites to find all candidate guide RNA sequences in your input FASTQ
  2. Optional Run crispr_sites to generate on-target and off-target files
  3. Run dashit-reads-filter to remove candidate gRNAs that have poor structure (homopolymer, dinucleotide repeats, hairpins), represent adapter sequences, hit off targets, or don’t hit on-targets
  4. Run optimize_guides to find the gRNAs that hit the largest number of reads in your input FASTQ

To see how your designed guides perform, score your reads

Scoring reads