Nanopore sequencing, as represented by Oxford Nanopore Technologies' MinION, is a promising technology for in situ life detection and for microbial monitoring including in support of human space ...exploration, due to its small size, low mass (~100 g) and low power (~1 W). Now ubiquitous on Earth and previously demonstrated on the International Space Station (ISS), nanopore sequencing involves translocation of DNA through a biological nanopore on timescales of milliseconds per base. Nanopore sequencing is now being done in both controlled lab settings as well as in diverse environments that include ground, air, and space vehicles. Future space missions may also utilize nanopore sequencing in reduced gravity environments, such as in the search for life on Mars (Earth-relative gravito-inertial acceleration (GIA)
= 0.378), or at icy moons such as Europa (
= 0.134) or Enceladus (
= 0.012). We confirm the ability to sequence at Mars as well as near Europa or Lunar (
= 0.166) and lower
levels, demonstrate the functionality of updated chemistry and sequencing protocols under parabolic flight, and reveal consistent performance across
level, during dynamic accelerations, and despite vibrations with significant power at translocation-relevant frequencies. Our work strengthens the use case for nanopore sequencing in dynamic environments on Earth and in space, including as part of the search for nucleic-acid based life beyond Earth.
All known life uses informational polymers based on nucleic acids. Future missions to Mars and Ocean Worlds such as Enceladus and/or Europa may target these or related polymers in the search for ...extant life beyond Earth. Nanopore-based devices represent a promising approach for sensing and characterizing these, and possibly other, biomarkers. Here we demonstrate low-input (200 pg)DNA sequencing, equivalent to extraction from 10 6 Bacillus subtilis spores at 5 percent extraction yield, using the Oxford Nanopore MinION in a thermal vacuum chamber under Mars-like temperature (−60°C), atmosphere (100% CO 2 ), and pressure (400 to 500 Pa). Current limits of detection correspond to 2 to 5 pg DNA. With additional advances in nucleic acid extraction and library preparation efficiency, a sequencing-based approach to life detection will be viable at cell densities representative of the most extreme Mars analog environments here on Earth.
Parabolic flights provide cost-effective, time-limited access to "weightless" or reduced gravity conditions, facilitating research and validation activities that complement infrequent and costly ...access to space. Although parabolic flights have been conducted for decades, reference acceleration profiles and processing methods are not widely available. Here we present a solution for collecting, analyzing, and classifying the altered gravity environments experienced during parabolic flights, which we validated during a Boeing 727-200F flight with 20 parabolas. All data and analysis code are freely available. Our solution can be integrated with diverse experimental designs, does not depend upon accelerometer orientation, and allows unsupervised classification of all phases of flight, providing a consistent and open-source approach to quantifying gravito-inertial accelerations (GIA), or
levels. As academic, governmental, and commercial use of space advances, data availability and validated processing methods will enable better planning, execution, and analysis of parabolic flight experiments, and thus facilitate future space activities.
This paper investigates the reliability of off the shelf Conductive Bridging Random Access Memory (CBRAM) chips under exposure to cold temperatures. In comparison to electrically erasable ...programmable read-only memory (EEPROM) or Flash memory, de-facto standards in spaceflight, CBRAM features radiation resistance, low power consumption, and high read and write speeds. In testing commercial grade and radiation resistant medical grade CBRAM chips, in biased (powered, actively used) and unbiased configurations, we show that: (a) unbiased chips exhibit no reading or writing errors after exposure to - 150°C; (b) nearly all (97%) of the 32 biased chips that were tested to - 50°C were still functional while (c) 75% of all 48 biased chips that were tested down to -75°C were functional. We detected no difference in performance between medical grade and non-medical grade chips, and every chip was fully functional once brought back to room temperature. With the ability to endure space radiation, operate on low power, and function with only minimal heating, CBRAM has potential as a memory solution on satellites or in space instruments.
Centaurs, minor planets with a semi-major axis between the orbits of Jupiter and Neptune (5–30 AU), are thought to be among the most diverse small bodies in the solar system. These important targets ...for future missions may have recently been Kuiper Belt Objects (KBOs), which are thought to be chemically and physically primitive remnants of the early solar system. While the Kuiper Belt spans distances of 30–50 AU, making direct observations difficult, Centaurs' proximity to the Earth and Sun make them more accessible targets for robotic missions. Thus, we outline a mission concept designed to reconnoiter 10199 Chariklo, the largest Centaur and smallest ringed body yet discovered. Named for a legendary Centaur tamer, the conceptual Camilla mission is designed to fit under the cost cap of the National Aeronautics and Space Administration (NASA) New Frontiers program, leveraging a conservative payload to support a foundational scientific investigation to these primitive bodies. Specifically, the single flyby encounter utilizes a combined high-resolution camera/VIS-IR mapping spectrometer, a sub-mm point spectrometer, and a UV mapping spectrometer. In addition, the mission concept utilizes a kinetic impactor, which would provide the first opportunity to sample the composition of potentially primitive subsurface material beyond Saturn, thus providing key insights into solar system origins. Such a flyby of the Chariklo system would provide a linchpin in the understanding of small body composition, evolution, and transport of materials in the solar system.
Display omitted
•Flyby concept for 10199 Chariklo, the largest Centaur and smallest ring system.•Opportunity to learn about Kuiper Belt Objects much closer to Earth.•Impactor would provide deepest yet subsurface sampling in the outer Solar System.•Mission concept fits well within NASA New Frontiers Program cost cap.•Mission concept may fit within NASA Discovery Program cost cap.
Future space instruments will explore increasingly complex questions about our universe, including the origin of life on Earth and the presence of life elsewhere. These instruments will likely ...integrate chemical and biological subsystems that will face unique challenges; existing protocols typically require non-stabilized components and manual handling. The Search for Extra-Terrestrial Genomes (SETG) instrument is being developed for in situ extraction and sequencing of nucleic acids as a biomarker of life on other planetary bodies. Such sequencing is being implemented using a nanopore-based device, the Oxford Nanopore Technologies (ONT) MinION; as such, it needs to integrate many benchtop-based protocols. Here we describe an automated testbed, designed and built to automate and rapidly prototype extraction, library preparation, and sequencing protocols that could be used in our instrument. The system is designed to be modular with respect to components, facilitating hardware and software modifications with minimal system impact, while also precise across multiple test runs, allowing for accurate evaluation of the impact of varying system inputs as well as exploration of system failure modes and potential solutions. We also present testing results from each of the three primary subsystems (extraction, library preparation, and sequencing) as well as a plan for and initial data on subsystem integration into an end-to-end system. The extraction subsystem is able to match or approach nucleic acid yields attained via manual testing for B. subtilis spores in water (~15 %) and spores in basalt (~ 12%). The library preparation subsystem can successfully prepare a library of E. coli DNA that can be identified after sequencing. The loading/sequencing subsystem has successfully automated sequencer loading, resulting in a sequencing run producing 1.4 billion bases after 1 day of sequencing from a pre-prepared sample. These testing results provide valuable data about the challenges of biological protocol automation, while directly informing future design decisions for SETG. In the process, the lessons learned from this milestone are relevant to the technological development of future planetary science instruments that take advantage of molecular biology techniques.
Due to meteoritic transfer between Earth and Mars, if life on Mars exists, it may be related to life on Earth and utilize nucleic acids as informational polymers. Thus, a Search for Extra-Terrestrial ...Genomes (SETG) could detect and sequence (deoxy)ribonucleic acids (DNA/RNA) utilized by any extant or recently dead life on Mars. The abiotic synthesis of common organic building blocks, such as nucleobases, sugars, and amino acids, in the solar nebula and potentially in diverse habitable environments could also bias a second genesis of life towards utilizing informational polymers similar to life as we know it. Here we build on prior work and describe the advancement of a SETG instrument to technology readiness level 4 through sample-to-sequence processing with limited manual handling. Another advance includes validation of nucleic acid extraction from Mars analogs at cell counts down to 10 4 per 50 mg sample, equivalent to a limit of detection of approximately 1 part per billion. In addition, we demonstrate that biological nanopore-based single molecule sequencing can be used to detect non-standard bases. Finally, we link sequence data to a statistical test to distinguish between any forward contamination and putative life beyond Earth. Nanopore-based sensing may ultimately enable characterization of non-standard polymers and other molecules, highlighting the potential for nanopore-based life detection and sequencing on Mars or other words such as the icy moons Enceladus or Europa.
Widespread synthesis of complex organics, including nucleobases and ribose precursors, occurred early in the history of the solar system in the solar nebula. These organics, delivered to multiple ...potentially habitable zones, may have biased the evolution of life towards utilization of similar informational polymers. Meteoritic exchange might also have produced shared ancestry, most plausible for Earth and Mars. To test this hypothesis, we are developing the Search for Extra-Terrestrial Genomes (SETG), a life detection instrument for in-situ isolation and sequencing of nucleic acids. Our mission focus area is astrobiology and the search for life beyond Earth. Our science goal for Mars is to search for related or unrelated nucleic acid-based life, particularly life that has the potential to interact with life on Earth; this may also inform sample selection for Mars Sample Return (MSR) and reduce the risks of false positives through the first in-situ measurement of forward contamination. Our science goal for Enceladus is to search for a second genesis based on nucleic acids in the plumes emanating from the South Polar Region. Life detection may also be possible in Europa orbit but the availability of a suitable plume is tenuous and it is a challenge for biological reagents to survive intense radiation there. Here we describe advancements in SETG geared towards in-situ sequencing during a future Mars mission, including extraction of nucleic acids coupled with proof of principle for in-situ single-molecule nanopore-based sequencing. We briefly describe plans to advance SETG from Technology Readiness Level 3 to 6 in preparation for future flight definition and show that under realistic assumptions, a sensitivity of parts per billion or better is feasible.
Parabolic flights provide cost-effective, time-limited access to "weightless" or reduced gravity conditions experienced in space or on planetary surfaces, e.g. the Moon or Mars. These flights ...facilitate fundamental research - from materials science to space biology - and testing/validation activities that support and complement infrequent and costly access to space. While parabolic flights have been conducted for decades, reference acceleration profiles and processing methods are not widely available - yet are critical for assessing the results of these activities. Here we present a method for collecting, analyzing, and classifying the altered gravity environments experienced during a parabolic flight. We validated this method using a commercially available accelerometer during a Boeing 727-200F flight with \(20\) parabolas. All data and analysis code are freely available. Our solution can be easily integrated with a variety of experimental designs, does not depend upon accelerometer orientation, and allows for unsupervised and repeatable classification of all phases of flight, providing a consistent and open-source approach to quantifying gravito-intertial accelerations (GIA), or \(g\) levels. As academic, governmental, and commercial use of space increases, data availability and validated processing methods will enable better planning, execution, and analysis of parabolic flight experiments, and thus, facilitate future space activities.
Thesis: S.M., Massachusetts Institute of Technology, Department of Aeronautics and Astronautics, 2018.
Cataloged from PDF version of thesis.
Includes bibliographical references (pages 75-78).
The ...development of any spaceflight instrument involves a systematic, iterative process of design, testing, and validation. This ensures that the system developed will meet the needs of stakeholders while minimizing costs and risks. Here, the needs for a life-detection instrument targeting nucleic acids are identified, the objectives for that instrument are determined, and system engineering analysis is used to demonstrate that a Search for Extra-Terrestrial Genomes instrument would fulfill those objectives and is feasible for a rover mission. Additionally, we show our design and build process for a testbed to rapidly prototype SETG components and subsystems, which has successfully automated nucleic acid extraction, sequencer loading, and parts of library preparation. We also experiment with thermal simulations and conduct a sequencing test at Martian conditions, using a custom built thermal vacuum chamber system. Finally, this thesis explores potential avenues for future development and identifies short term and long term engineering goals that would assist the SETG team in developing an instrument prototype.
by Srinivasa Aditya Bhattaru.
S.M.
PDF
