Mixing Illumina and PacBio Data for Genome Finishing
PacBio machines give long reads, but the sequences are very noisy (error rate 1 in 10). It may be possible to improve the quality of PacBio sequences by multiple rounds of resequencing, but a new (and cheaper) cleanup alternative is emerging that uses short high-quality Illumina reads. We earlier discussed how the method works based on slides from Michael Schatz, but now the readers can find more information from two recently published papers.
1. Finished bacterial genomes from shotgun sequence data
A paper published in Genome Research from Broad Institute ALLPATHS team discussed how they automated the expensive finishing step of bacterial genomes by combining data from two technologies.
By applying a new laboratory design and new assembly algorithm to sixteen samples, we demonstrate that assemblies exceeding finished quality can be obtained from whole-genome shotgun data and automated computation. Cost and time requirements are thus dramatically reduced.
2. Hybrid error correction and de novo assembly of single-molecule sequencing reads
This paper is from Michael Schatz’s group.
We introduce a correction algorithm and assembly strategy that uses short, high-fidelity sequences to correct the error in single-molecule sequences. We demonstrate the utility of this approach on reads generated by a PacBio RS instrument from phage, prokaryotic and eukaryotic whole genomes, including the previously unsequenced genome of the parrot Melopsittacus undulatus, as well as for RNA-Seq reads of the corn (Zea mays) transcriptome. Our long-read correction achieves >99.9% base-call accuracy, leading to substantially better assemblies than current sequencing strategies: in the best example, the median contig size was quintupled relative to high-coverage, second-generation assemblies.
Readers may also look at PacBioToCA software from the same group.
Needless to say, neither paper answers, or can possibly answer, an important question regarding PacBio in many people’s mind. Can any algorithm turn the slope of this curve upward?
