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Readfish enables targeted nanopore sequencing of gigabase-sized genomes

Nanopore sequencers can be used to selectively sequence certain DNA molecules in a pool by reversing the voltage across individual nanopores to reject specific sequences, enabling enrichment and depletion to address biological questions. Previously, we achieved this using dynamic time warping to map...

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Detalles Bibliográficos
Autores principales: Payne, Alexander, Holmes, Nadine, Clarke, Thomas, Munro, Rory, Debebe, Bisrat J, Loose, Matthew
Formato: Online Artículo Texto
Lenguaje:English
Publicado: 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7610616/
https://www.ncbi.nlm.nih.gov/pubmed/33257864
http://dx.doi.org/10.1038/s41587-020-00746-x
Descripción
Sumario:Nanopore sequencers can be used to selectively sequence certain DNA molecules in a pool by reversing the voltage across individual nanopores to reject specific sequences, enabling enrichment and depletion to address biological questions. Previously, we achieved this using dynamic time warping to map the signal to a reference genome, but the method required substantial computational resources and did not scale to gigabase-sized references. Here we overcome this limitation by using GPU base calling. We show enrichment of specific chromosomes from the human genome and of low-abundance organisms in mixed populations without a priori knowledge of sample composition. Finally, we enrich targeted panels comprising 25,600 exons from 10,000 human genes and 717 genes implicated in cancer, identifying PML-RARA fusions in the NB4 cell line in <15 hours sequencing. These methods can be used to efficiently screen any target panel of genes without specialised sample preparation using any computer and suitable GPU. Our toolkit, readfish, is available at https://www.github.com/looselab/readfish.