182
views
0
recommends
+1 Recommend
0 collections
    0
    shares
      • Record: found
      • Abstract: found
      • Article: not found

      Improved data analysis for the MinION nanopore sequencer

      research-article

      Read this article at

      ScienceOpenPublisherPMC
      Bookmark
          There is no author summary for this article yet. Authors can add summaries to their articles on ScienceOpen to make them more accessible to a non-specialist audience.

          Abstract

          The Oxford Nanopore MinION sequences individual DNA molecules using an array of pores that read nucleotide identities based on ionic current steps. We evaluated and optimized MinION performance using M13 genomic dsDNA. Using expectation-maximization (EM) we obtained robust maximum likelihood (ML) estimates for read insertion, deletion and substitution error rates (4.9%, 7.8%, and 5.1% respectively). We found that 99% of high-quality ‘2D’ MinION reads mapped to reference at a mean identity of 85%. We present a MinION-tailored tool for single nucleotide variant (SNV) detection that uses ML parameter estimates and marginalization over many possible read alignments to achieve precision and recall of up to 99%. By pairing our high-confidence alignment strategy with long MinION reads, we resolved the copy number for a cancer/testis gene family (CT47) within an unresolved region of human chromosome Xq24.

          Related collections

          Most cited references27

          • Record: found
          • Abstract: not found
          • Book: not found

          Biological sequence analysis

            Bookmark
            • Record: found
            • Abstract: found
            • Article: not found

            MinION nanopore sequencing identifies the position and structure of a bacterial antibiotic resistance island.

            Short-read, high-throughput sequencing technology cannot identify the chromosomal position of repetitive insertion sequences that typically flank horizontally acquired genes such as bacterial virulence genes and antibiotic resistance genes. The MinION nanopore sequencer can produce long sequencing reads on a device similar in size to a USB memory stick. Here we apply a MinION sequencer to resolve the structure and chromosomal insertion site of a composite antibiotic resistance island in Salmonella Typhi Haplotype 58. Nanopore sequencing data from a single 18-h run was used to create a scaffold for an assembly generated from short-read Illumina data. Our results demonstrate the potential of the MinION device in clinical laboratories to fully characterize the epidemic spread of bacterial pathogens.
              Bookmark
              • Record: found
              • Abstract: found
              • Article: not found

              Automated Forward and Reverse Ratcheting of DNA in a Nanopore at Five Angstrom Precision1

              Single-molecule techniques have been developed for commercial DNA sequencing 1,2 . One emerging strategy uses a nanopore to analyze DNA molecules as they are driven electrophoretically in single file order past a sensor 3-5 . However, uncontrolled DNA strand electrophoresis through nanopores is too fast for accurate base reads 6 . A proposed solution would employ processive enzymes to deliver DNA through the pore at a slower average rate 7 . Here, we describe forward and reverse ratcheting of DNA templates through the α–hemolysin (α-HL) nanopore controlled by wild-type phi29 DNA polymerase (phi29 DNAP). DNA strands were examined in single file order at one nucleotide spatial precision in real time. The registry error probability (either an insertion or deletion during one pass along a template strand) ranged from 10% to 24.5% absent optimization. This general strategy facilitates multiple reads of individual template strands and is transferrable to other nanopore devices for implementation of DNA sequence analysis.
                Bookmark

                Author and article information

                Journal
                101215604
                32338
                Nat Methods
                Nat. Methods
                Nature methods
                1548-7091
                1548-7105
                6 June 2016
                16 February 2015
                April 2015
                14 June 2016
                : 12
                : 4
                : 351-356
                Affiliations
                [1 ]University of California Santa Cruz Genomics Institute, Santa Cruz, CA USA
                [2 ]Department of Biomolecular Engineering, University of California, Santa Cruz, CA USA
                Author notes
                Correspondence: bioinformatics, benedict@ 123456soe.ucsc.edu ; nanopore technology, makeson@ 123456soe.ucsc.edu
                Article
                NIHMS658412
                10.1038/nmeth.3290
                4907500
                25686389
                009ff1f4-b4a6-4cf8-832c-3c7a9a024701

                Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use: http://www.nature.com/authors/editorial_policies/license.html#terms

                History
                Categories
                Article

                Life sciences
                Life sciences

                Comments

                Comment on this article