FLASH: a next-generation CRISPR diagnostic for multiplexed detection of antimicrobial resistance sequences.
; Mourani, Peter M; Calfee, Carolyn S; Neff, Norma F; Chow, Eric D; Kim, Peter S; Greenhouse, Bryan; DeRisi, Joseph L; and Crawford, Emily D
(2019)
FLASH: a next-generation CRISPR diagnostic for multiplexed detection of antimicrobial resistance sequences.
Nucleic acids research, 47 (14).
e83-.
ISSN 0305-1048
DOI: 10.1093/nar/gkz418
The growing prevalence of deadly microbes with resistance to previously life-saving drug therapies is a dire threat to human health. Detection of low abundance pathogen sequences remains a challenge for metagenomic Next Generation Sequencing (NGS). We introduce FLASH (Finding Low Abundance Sequences by Hybridization), a next-generation CRISPR/Cas9 diagnostic method that takes advantage of the efficiency, specificity and flexibility of Cas9 to enrich for a programmed set of sequences. FLASH-NGS achieves up to 5 orders of magnitude of enrichment and sub-attomolar gene detection with minimal background. We provide an open-source software tool (FLASHit) for guide RNA design. Here we applied it to detection of antimicrobial resistance genes in respiratory fluid and dried blood spots, but FLASH-NGS is applicable to all areas that rely on multiplex PCR.
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