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A universal, genome-wide guide finder for CRISPR/Cas9 targeting in microbial genomes

Michelle Spoto, Elizabeth Fleming, Julia Oh
doi: https://doi.org/10.1101/194241
Michelle Spoto
1The Jackson Laboratory for Genomic Medicine, 10 Discovery Drive Farmington, CT, , ,
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  • For correspondence: Michelle.Spoto@jax.org Elizabeth.Fleming@jax.org Julia.Oh@jax.org
Elizabeth Fleming
1The Jackson Laboratory for Genomic Medicine, 10 Discovery Drive Farmington, CT, , ,
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  • For correspondence: Michelle.Spoto@jax.org Elizabeth.Fleming@jax.org Julia.Oh@jax.org
Julia Oh
1The Jackson Laboratory for Genomic Medicine, 10 Discovery Drive Farmington, CT, , ,
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  • For correspondence: Julia.Oh@jax.org Michelle.Spoto@jax.org Elizabeth.Fleming@jax.org Julia.Oh@jax.org
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Abstract

Background The CRISPR/Cas system has significant potential to facilitate gene editing in a variety of bacterial species. CRISPR interference (CRISPRi) and CRISPR activation (CRISPRa) represent modifications of the CRISPR/Cas9 system utilizing a catalytically inactive Cas9 protein for transcription repression or activation, respectively. While CRISPRi and CRISPRa have tremendous potential to systematically investigate gene function in bacteria, no pan-bacterial, genome-wide tools exist for guide discovery. We have created Guide Finder: a customizable, user-friendly program that can design guides for any annotated bacterial genome.

Results Guide Finder designs guides from NGG PAM sites for any number of genes using an annotated genome and fasta file input by the user. Guides are filtered according to user-defined design parameters and removed if they contain any off-target matches. Iteration with lowered parameter thresholds allows the program to design guides for genes that did not produce guides with the more stringent parameters, a feature unique to Guide Finder. Guide Finder has been tested on a variety of diverse bacterial genomes, on average finding guides for 95% of genes. Moreover, guides designed by the program are functionally useful—focusing on CRISPRi as a potential application—as demonstrated by essential gene knockdown in two staphylococcal species.

Conclusions Through the large-scale generation of guides, this open-access software will improve accessibility to CRISPR/Cas studies for a variety of bacterial species.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. All rights reserved. No reuse allowed without permission.
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Posted September 27, 2017.
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A universal, genome-wide guide finder for CRISPR/Cas9 targeting in microbial genomes
Michelle Spoto, Elizabeth Fleming, Julia Oh
bioRxiv 194241; doi: https://doi.org/10.1101/194241
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A universal, genome-wide guide finder for CRISPR/Cas9 targeting in microbial genomes
Michelle Spoto, Elizabeth Fleming, Julia Oh
bioRxiv 194241; doi: https://doi.org/10.1101/194241

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