Nanopore sequencing in microgravity

Abstract

Rapid DNA sequencing and analysis has been a long-sought goal in remote research and point-of-care medicine. In microgravity, DNA sequencing can facilitate novel astrobiological research and close monitoring of crew health, but spaceflight places stringent restrictions on the mass and volume of instruments, crew operation time, and instrument functionality. The recent emergence of portable, nanopore-based tools with streamlined sample preparation protocols finally enables DNA sequencing on missions in microgravity. As a first step toward sequencing in space and aboard the International Space Station (ISS), we tested the Oxford Nanopore Technologies MinION during a parabolic flight to understand the effects of variable gravity on the instrument and data. In a successful proof-of-principle experiment, we found that the instrument generated DNA reads over the course of the flight, including the first ever sequenced in microgravity, and additional reads measured after the flight concluded its parabolas. Here we detail modifications to the sample-loading procedures to facilitate nanopore sequencing aboard the ISS and in other microgravity environments. We also evaluate existing analysis methods and outline two new approaches, the first based on a wave-fingerprint method and the second on entropy signal mapping. Computationally light analysis methods offer the potential for in situ species identification, but are limited by the error profiles (stays, skips, and mismatches) of older nanopore data. Higher accuracies attainable with modified sample processing methods and the latest version of flow cells will further enable the use of nanopore sequencers for diagnostics and research in space.

Genetic and epigenetic heterogeneity and the impact on cancer relapse

“…Moreover, new single-base resolution methods are also emerging, which can leverage regional, phased, and long-range information from differentially methylated regions, single molecule sequencing, and new nanopore-based methods [28–30]. Notabl, y these techniques are already leading to new combined therapies and measure in leukemia patients with specifical mutations, like DNMT3A and TET mutations [31–32].…”

Towards a genomics-informed, real-time, global pathogen surveillance system

“…Importantly, the MinION has been used in field situations, including in diagnostic tent labora tories during the Ebola epidemic 22,23 and in a roving busbased mobile laboratory in Brazil as part of the ZiBRA project 3,24 . Others have taken the MinION to more extreme environments where even the smallest traditional bench-top sequencer could not go, including the Arctic 25 and Antarctic 26 , a deep mine 27 and zero gravity aboard the reduced-gravity aircraft (nicknamed the 'Vomit Comet') 28 and the International Space Station 29 . However, this technology is not yet a panacea; remaining challenges include high DNA or RNA input requirements (currently hundreds of nanograms), which often necessitate PCR-based amplification approaches; a flow cell cost of $500, keeping the cost per sample high despite multiplexing approaches; and high error rates, which require that genomes are sequenced to high coverage for single nucleotide polymorphism-based analysis and analysed at the signal level.…”

Nucleic acid extraction and sequencing from low-biomass synthetic Mars analog soils for in situ life detection

“…Prior studies have utilized OmniLyse, which is Purelyse® without extraction buffers, to lyse cells in an RNA extraction module aboard the International Space Station (Parra et al, 2017). Long-read sequencing is then conducted using the Oxford Nanopore Technologies (ONT) MinION which sequences nucleic acids via ionic current monitoring (Lu et al, 2016) and has been validated to sequence DNA in microgravity (Castro-Wallace et al, 2017;McIntyre et al, 2016), Lunar and Mars gravity (Carr/Zuber in prep 2018), and under simulated Mars temperature and pressure (Carr in prep 2018).…”