Recovering individual haplotypes and a contiguous genome assembly from pooled long read sequencing of the diamondback moth (Lepidoptera: Plutellidae)

Whiteford, S.; van't Hof, A. E.; Krishna, R.; Marubbi, T.; Widdison, S.; Saccheri, I. J.; Guest, M.; Morrison, N. I.; Darby, A. C.

doi:10.1101/867879

bioRxiv · 10.1101/867879

Recovering individual haplotypes and a contiguous genome assembly from pooled long read sequencing of the diamondback moth (Lepidoptera: Plutellidae)

Whiteford, S.·van't Hof, A. E.·Krishna, R.·Marubbi, T.·Widdison, S.·Saccheri, I. J.·Guest, M.·Morrison, N. I.·Darby, A. C.

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Abstract

BackgroundRecent advances in genomics have addressed the challenge that divergent haplotypes pose to the reconstruction of haploid genomes. However for many organisms, the sequencing of either field-caught individuals or a pool of heterogeneous individuals is still the only practical option. Here we present methodological approaches to achieve three outcomes from pooled long read sequencing: the generation of a contiguous haploid reference sequence, the sequences of heterozygous haplotypes; and reconstructed genomic sequences of individuals related to the pooled material. ResultsPacBio long read sequencing, Dovetail Hi-C scaffolding and linkage map integration yielded a haploid chromosome-level assembly for the diamondback moth (Plutella xylostella), a global pest of Brassica crops, from a pool of related individuals. The final assembly consisted of 573 scaffolds, with a total assembly size of 343.6Mbp a scaffold N50 value of 11.3Mbp (limited by chromosome size) and a maximum scaffold size of 14.4Mbp. This assembly was then integrated with an existing RAD-seq linkage map, anchoring 95% of the assembled sequence to defined chromosomal positions. ConclusionsWe describe an approach to resolve divergent haplotype sequences and describe multiple validation approaches. We also reconstruct individual genomes from pooled long-reads, by applying a recently developed k-mer binning method.

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Whiteford, S., van't Hof, A. E., Krishna, R., Marubbi, T., Widdison, S., Saccheri, I. J., Guest, M., Morrison, N. I., Darby, A. C.. 2019-12-06. Recovering individual haplotypes and a contiguous genome assembly from pooled long read sequencing of the diamondback moth (Lepidoptera: Plutellidae). https://doi.org/10.1101/867879

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