A new paper on bacterial genome assembly titled “A single chromosome assembly of Bacteroides fragilis strain BE1 from Illumina and MinION nanopore sequencing data” has been submitted to biorxiv. The senior author is Mick Watson, an academia-based vocal promoter of the company. Therefore the paper likely shows the state-of-the-art of the technology.
The most puzzling observation one can make from the paper is the discrepancy between the claimed and the actual error rate of nanopore sequences. We recently had this discussion with David Eccles, who wrote in the comment section - “The median error per base was approximately 7% for this run (i.e. 93% accuracy)”. Similar claims of 92%-93% accuracy had been made by other Nanopore promoters. If that is true, the obvious follow-up question would be - why does Mick Watson (and many others) need to use Illumina reads to assemble bacterial genomes? For Pacbio reads with 85% accuracy, one can easily assemble bacterial genomes from Pacbio-only.
The answer lies in two ‘details’. (i) For Pacbio reads, 85% accuracy is before processing, whereas Oxford nanopore reads appear to go through rounds of processing to get to 92%. I pointed that out to David Eccles in the comment section. (ii) The second and more important point is that the error distribution of Pacbio reads is completely random, thus making compensation through high coverage possible. We are not sure what the error distribution of Oxford nanopore reads is. It is possibly not random given that they continue to need support from Illumina sequences for genome assembly.
The hybrid assembly argument vouched in the following abstract was the status quo for Pacbio one year back, but based on my recent conversation with the members of Pacbio community, those researchers are increasingly moving away from using any Illumina read. Apparently, those short reads seem to complicate the assembly process instead of helping. They were useful, when Pacbio reads were expensive, but will not be any more.
Second and third generation sequencing technologies have revolutionised bacterial genomics. Short-read Illumina reads result in cheap but fragmented assemblies, whereas longer reads are more expensive but result in more complete genomes. The Oxford Nanopore MinION device is a revolutionary mobile sequencer that can produce thousands of long, single molecule reads.
We sequenced Bacteroides fragilis strain BE1 using both the Illumina MiSeq and Oxford Nanopore MinION platforms. We were able to assemble a single chromosome of 5.18 Mb, with no gaps, using publicly available software and commodity computing hardware. We identified gene rearrangements and the state of invertible promoters in the strain.
The single chromosome assembly of Bacteroides fragilis strain BE1 was achieved using only modest amounts of data, publicly available software and commodity computing hardware. This combination of technologies offers the possibility of ultra-cheap, high quality, finished bacterial genomes.
Sam Minot pointed out Jared Simpson’s work on E. coli assembly from Oxford nanopore only, which we covered here-
It is puzzling that despite that earlier success, others are not adopting similar approach, given that assembling from Oxford nanopore only would be the biggest selling point for the company. It does not make sense to talk about carrying USB-sized sequencer to the field, if one also has to carry a large Illumina instrument. There are two possible explanations for Mick Watson’s not using Simpson’s algorithm/program - (i) he is not aware of it, (ii) he is unable to make it work.
Other helpful twitter comment -