Abstract
SummaryNanopore sequencing is an increasingly powerful tool for genomics. Recently, computational advances have allowed nanopores to sequence in a targeted fashion; as the sequencer emits data, software can analyze the data in real time and signal the sequencer to eject “nontarget” DNA molecules. We present a novel method called SPUMONI, which enables rapid and accurate targeted sequencing using efficient pan-genome indexes. SPUMONI uses a compressed index to rapidly generate exact or approximate matching statistics in a streaming fashion. When used to target a specific strain in a mock community, SPUMONI has similar accuracy as minimap2 when both are run against an index containing many strains per species. However SPUMONI is 12 times faster than minimap2. SPUMONI's index and peak memory footprint are also 16 to 4 times smaller than those of minimap2, respectively. This could enable accurate targeted sequencing even when the targeted strains have not necessarily been sequenced or assembled previously.
Highlights
Nanopore sequencing instruments have steadily improved in usability, speed, and accuracy
To determine whether the matching statistics (MSs) are long enough to indicate an approximate match, SPUMONI compares the observed distribution of MSs – calculated with respect to a ‘‘positive index’’ containing the target sequences – with those obtained from a ‘‘null index’’ containing the reverse of the sequences from the positive index
As soon as SPUMONI can confidently determine the distributions of MSs from the positive and null indexes are different – possibly having seen only a prefix of the read’s full sequence – it can conclude that the read is among the targets in the positive index
Summary
Nanopore sequencing instruments have steadily improved in usability, speed, and accuracy. Nanopore sequencing is convenient and flexible; nanopores are readily used outside of laboratories, for example, for analyzing biological species in a human or natural environment with the goal of detecting pathogens or contaminants. They can be used for several assays, including DNA sequencing, direct RNA sequencing, and the detection of a variety of epigenetic modifications. Nanopore sequencing has the unique capability where the control software can potentially signal to the sequencer that it should eject the DNA molecule currently in a pore. Many such pores – up to 512 per MinION flowcell – are in simultaneous operation; the system can sequence in a targeted manner only as long as the software making ejection decisions can keep up with the aggregate rate of sequencing
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