Nanopore sequencing and assembly of a human genome with ultra-long reads

Nanopore sequencing is a promising technique for genome sequencing due to its portability, ability to sequence long reads from single molecules, and to simultaneously assay DNA methylation. However until recently nanopore sequencing has been mainly applied to small genomes, in part due to the limited output attainable. Here, we present nanopore sequencing and assembly of the GM12878 Utah/Ceph human reference genome generated using the Oxford Nanopore MinION and R9.4 version chemistry. We generated 91.2 Gb of sequence data (~30× theoretical coverage) from 39 flowcells. De novo assembly yielded a highly complete and contiguous assembly (NG50 ~3 Mb). We observed considerable variability in homopolymeric tract resolution between different basecallers. The data permitted sensitive detection of both large structural variants and epigenetic modifications. Further, we developed a new approach exploiting the long-read capability of this system and found that adding an additional 5×-coverage of 'ultra-long' reads more than doubled the assembly contiguity (NG50 ~6.4 Mb). These reads enabled complete phasing across the MHC in a single contig, determination of telomere repeat length and closure of gaps in the existing reference assembly. Modelling the repeat structure of the human genome predicts extraordinarily contiguous assemblies may be possible using nanopore reads alone.