The use of Oxford Nanopore native barcoding for complete genome assembly

Background: The Oxford Nanopore Technologies MinION(TM) is a mobile DNA sequencer that can produce long read sequences with a short turn-around time. Here we report the first demonstration of single contig genome assembly using Oxford Nanopore native barcoding when applied to a multiplexed library of 12 samples and combined with existing Illumina short-read data. This paves the way for the closure of multiple bacterial genomes from a single MinION(TM) sequencing run, given the availability of existing short-read data. The strain we used, MHO_001, represents the important community-acquired methicillin resistant Staphylococcus aureus lineage USA300.
Findings: Using a hybrid assembly of existing short read and barcoded long read sequences from multiplexed data, we completed a genome of the S. aureus USA300 strain MHO_001. The long-read data represented only ∼5-10% of an average MinION(TM) run (∼7x genomic coverage), but, using standard tools, this was sufficient to complete the circular chromosome of S. aureus strain MHO_001 (2.86 Mb) and two complete plasmids (27 Kb and 3 Kb). Minor differences were noted when compared to USA300 reference genome, USA300_FPR3757, including the translocation, loss and gain of mobile genetic elements.
Conclusion: Here we demonstrate that MinION(TM) reads, multiplexed using native barcoding, can be used in combination with short-read data, to fully complete a bacterial genome. The ability to complete multiple genomes, for which short-read data is already available, from a single MinION(TM) run is set to impact on our understanding of accessory genome content, plasmid diversity and genome rearrangements.