Depletion of Abundant Sequences by Hybridization (DASH): Using Cas9 to remove unwanted highabundance species in sequencing libraries and molecular counting applications
Gu W*, Crawford ED*, O’Donovan BD, Wilson MR, Chow ED, Retallack H, DeRisi JL (*These authors contributed equally)BioRxiv, 2015 and Genome Biology, 2016Abstract: Background:
With widespread adoption of next-generation sequencing (NGS) technologies, the need
has arisen for a broadly applicable method to remove unwanted high-abundance species prior to
sequencing. We introduce DASH (Depletion of Abundant Sequences by Hybridization), a facile
technique for targeted depletion of undesired sequences.
Sequencing libraries are DASHed with recombinant Cas9 protein complexed with a library of
single guide RNAs (sgRNAs) programmed to target unwanted species for cleavage, thus preventing
them from consuming sequencing space. We demonstrate up to 99% reduction of mitochondrial
ribosomal RNA (rRNA) in HeLa cells, and enrichment of pathogen sequences up to 4-fold in
metagenomic samples from patients with infectious diseases. Similarly, we demonstrate the utility of
DASH in the context of cancer diagnostics by significantly increasing the detectable fraction of KRAS
mutant sequences over the predominant wild-type allele.
This simple single-tube method is reprogrammable for virtually any sample type to
increase sequencing yield without additional cost.