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  • Oxford Nanopore Technologies

    Hadas Keren-Shaul Advanced Sequencing Technologies (Sandbox), LSCF Feinberg June 2018

    LSCF

  • Nanopore sequencing – Long reads

    Long read sequencing technology offers simplified and less ambiguous genome assembly

  • “Analysis of any living thing, by any person, in any environment”

    MinION – The first and only nanopore DNA sequencer

    • A portable, real-time, long-read, low-cost device

    • Designed to bring easy biological analyses to scientific research, education or a range of real-world applications such as disease/pathogen surveillance, environmental monitoring, food-chain surveillance, self-quantification or even microgravity biology

    • Every flow cell - 10-20 Gb of DNA sequence data

  • MinION sequencing device

    Lu et al., Review: Oxford nanopore MinION sequencing and genome assembly, Genomics Proteomics Bioinformatics, 2016

  • What is a Nanopore?

    • A very small hole – nano-scale

    • Oxford nanopore passes an ionic current through nanopores and measures the changes as biological molecules pass through the nanopore or near it

    • Holes can be created by protein puncturing membranes (biological nanopores) or in solid materials (solid-state nanopores)

  • How does it work?

    https://nanoporetech.com/how-it-works

    • A strand of DNA is passed through a nanopore

    • The current is changed as the bases G, A, T and C pass through the pore in different combinations

    https://nanoporetech.com/how-it-works

  • Direct molecular analysis

    • A direct electronic analysis of the target molecule – native analysis

    • Sequencing native DNA (and RNA) avoids bias (no amplification)

  • Rapid sequencing kit

    • Input – extracted gDNA

    • Time – 10 minutes

    • A transposase which simultaneously cleaves template molecules and attaches tags to the cleaved ends

  • cDNA PCR sequencing kit

    • Input – 50 ng poly(A) RNA

    • RT and PCR

  • Direct RNA sequencing kit

    • Input – 500 ng poly(A) RNA

    • No RT or PCR bias

    • Useful for transcripts that are difficult to reverse transcribe

  • The Nanopore sequencing workflow

    • Real Time data: no fixed run time allows on-demand sequencing and real time adjustment

  • Why MinION?

  • Applications

  • Making nanopore scalable

    A core sensing unit of a nanopore set in an arrayed sensor chip

  • PacBio vs. Oxford Nanopore

    • PacBio – Sequence by synthesis

    • Nanopore – monitoring the transit of a DNA molecule through a hole and measuring variations in electric current

  • PacBio vs. Oxford Nanopore

    • PacBio has overall better data quality

    • ONT provides higher yield per flow cell

  • Thank you for listening