SETG: Search for Extra-Terrestrial Genomes

Performance Period : May 2015 to April 2021

Faculty Investigator : Prof. Maria T. Zuber (PI, MIT); Prof. Gary Ruvkun (Co-PI, MGH), Prof. Christopher E. Carr (Science PI, MIT/Georgia Tech)

Project Status : Completed

Collaborators: Robert Doebler (Clarmont Biosolutions LLC), Michael Finney (self) and numerous other contributors, students, and staff.

  • High School Interns: Jonathan Borowsky
  • Undergraduate Researchers: Katherine M. Hahn, Ethan Hammons, Daniel Ledesma, Delson Faria Dasilva, Alex Choi, Carson Smith, Madeline Burgoyne, Jane Fessenden, Alex Smith
  • Staff: Julie Hachey, Kendall Saboda
  • Postdocs: Alexandra Pontefract, Jacopo Tani

The Search for Extra-Terrestrial Genomes (SETG) will search for nucleic acid-based life beyond Earth using single-molecule sequencing. Our MATISSE effort focused on developing an integrated instrument from technology readiness level (TRL) 3 to TRL 6 (sample in/sequence out under simulated Mars temperature/pressure) and assessing the limit of detection (LOD). We were able to advance component and/or subsystem TRLs as high as 6 in some cases, and achieved an overall TRL of 4 and partway to TRL5, therefore the current system TRL stands at TRL 4. At Georgia Tech in the science PI’s lab, work continues on the next phase of SETG, currently supported by faculty startup funds. This payload, Biological Exploration Payload 2 (BioX2) will integrate many aspects of SETG into an automated payload deployed on the International Space Station with a planned launch of October 2022. BioX2 will elevate the TRL level of several key components and subsystems in preparation for future flight definition, for example as a CubeSat demonstrator, and ultimately as part of an in-situ life detection mission. Key accomplishments and outcomes:

  • Enabled training of 17 students and/or staff and supported transition to tenure track or research faculty positions for 3 project members.
  • Supported 15 refereed conference or journal papers, indicated in bold as P1 to P15 as listed in the Publications section.
  • Synthesized Mars analog regolith samples doped with B. subtilis spores (P4) and validated a wide range of counting methods, identifying viable spore assay and/or Tunable Resistive Pulse Sensing (TRPS) as gold standards for quantifying spore titer.
  • Validated extraction procedure to meet requirements of 5% extraction yield (DNA out/DNA in) across all analog samples (P4, P12).
  • Developed procedure to count individual genomes using digital droplet PCR to facilitate evaluation of extraction efficiency at low biomass (P12)
  • Evaluated two single molecule nanopore sequencing technologies, and advanced strand sequencing (MinION) sequencing capabilities by:
    • detecting non-standard astrobiologically-relevant inosine bases (P3),
    • demonstrating a limit of detection of ~1 ppb for SETG using 2 picograms of DNA, orders of magnitude below the nominal input DNA requirement, by developing CarrierSeq (P6),
    • demonstrated a negligible false positive rate (P7),
    • performed sequencing under Mars-like conditions (P11), and
    • demonstrated robustness of nanopore sequencing to vibration and g-level (P13)
  • Automated nucleic acid extraction and demonstrated a TRL4 system through sample to sequence with limited manual handling (P3).
  • Developed a pre-TRL 5 testbed that facilitated risk reduction activities and demonstrated automation of all critical SETG steps (P10).
  • Designed and partially implemented a TRL5 SETG system.
  • Deployed one no-cost payload to the ISS to validate SETG reagents (BioX1).
  • Designed a second payload that will validate SETG hardware designs and advance SETG TRL through on-orbit operation (BioX2).

Funding: NASA Award # NNX15AF85G to Maria T. Zuber (MIT), with earlier support via NASA’s ASTID program (NNX08AX15G, NNG05GK27G) and associated fieldwork supported by NASA Awards NNX09AO76G and NNX12AM83G to Gary Ruvkun (MGH).

Publications (9 journal, 6 refereed conference papers):

P15. Carr CE. Resolving the History of Life on Earth by Seeking Life As We Know It on Mars. Astrobiology Jul 2022, 888-888, https://doi.org/10.1089/ast.2021.0043. Preprint available at: arXiv:2102.02362 https://arxiv.org/abs/2102.02362

P14. Mojarro, A, Jin, L, Szostak, JW, Head, JW, Zuber, MT. In search of the RNA world on Mars. Geobiology. 2021; 19: 307– 321. https://doi.org/10.1111/gbi.12433

P13. Carr CE, Bryan NC, Saboda K, Bhattaru SA, Ruvkun G, Zuber MT. Nanopore Sequencing at Mars, Europa, and Microgravity Conditions. npj Microgravity 6, Article number: 24 (2020). Preprint available at: https://www.biorxiv.org/content/10.1101/2020.01.09.899716v1. Data: https://osf.io/n6krq/ and https://osf.io/nk2w4/; Code: https://github.com/CarrCE/zerogseq

P12. A. Mojarro, J. Hachey, R. Bailey, M. Brown, R. Doebler, G. Ruvkun, M. T. Zuber, C. E. Carr. Nucleic Acid Extraction and Sequencing from Low-Biomass Synthetic Mars Analog Soils for In Situ Life Detection. Astrobiology 19(9) (2019) https://doi.org/10.1089/ast.2018.1929. Preprint: bioRxiv 358218 https://doi.org/10.1101/358218

P11. CE Carr, K Saboda, A Mojarro, J Hachey, SA Bhattaru, G Ruvkun, MT Zuber. Nucleic Acid Sequencing Under Mars-Like Conditions. Aerospace Conference, 2019 IEEE. March 2-9, Big Sky, MT, USA. Session 2.07 In Situ Instruments for Landed Surface Exploration, Orbiters and Flybys. Paper 2860. doi:10.1109/AERO.2019.8741543

P10. SA Bhattaru, J Tani, K Saboda, J Borowsky, G Ruvkun, MT Zuber, CE Carr. Development of a Nucleic Acid-Based Life Detection Instrument Testbed. Aerospace Conference, 2019 IEEE. March 2-9, Big Sky, MT. Session 2.07 In Situ Instruments for Landed Surface Exploration, Orbiters and Flybys. Paper 2181. doi:10.1109/AERO.2019.8742193

P9. K Saboda, R Moeller, CE Carr. Enabling Measurement of Darwinian Evolution in Space. Aerospace Conference, 2019 IEEE. March 2-9, Big Sky, MT, USA. Session 2.07 In Situ Instruments for Landed Surface Exploration, Orbiters and Flybys. Paper 2077. doi: 10.1109/AERO.2019.8742164

P8. Carr CE, Bryan NC, Saboda K, Bhattaru SA, Ruvkun G, Zuber MT. Acceleration Profiles and Processing Methods for Parabolic Flight. npj Microgravity 4, Article number: 14 (2018) https://doi.org/10.1038/s41526-018-0050-3. Preprint: arXiv:1712.05737; code: https://github.com/CarrCE/zerog; data: https://osf.io/nk2w4/

P7. A. Pontefract, J. Hachey, M. T. Zuber, G. Ruvkun, C. E. Carr. Sequencing Nothing: Exploring Failure Modes of Nanopore Sensing and Implications for Life Detection. Life Sciences in Space Research 2018 Aug;18:80-86. doi: 10.1016/j.lssr.2018.05.004. Epub 2018 May 16.

P6. Mojarro A, Hachey J, Ruvkun G, Zuber MT, Carr CE. CarrierSeq: a sequence analysis workflow for low-input nanopore sequencing. BMC Bioinformatics, 2018, 19:108, doi: 10.1186/s12859-018-2124-3 Preprint available at: bioRxiv https://doi.org/10.1101/175281; code: https://github.com/amojarro/carrierseq; data: https://doi.org/10.6084/m9.figshare.5471824.v1

P5. Pontefract A, Zhu TF, Walker VK, Rowedder H, Lui C, Zuber MT, Ruvkun G, Carr CE. Microbial Diversity in a Hypersaline Sulfate Lake: An Analog of Ancient Mars. Frontiers in Microbiology, 26 Sept 2017 doi: 10.3389/fmicb.2017.01819 PMID 29018418 OpenAccess

P4. Mojarro A, Ruvkun G, Zuber MT, Carr CE. Nucleic acid extraction from synthetic Mars analog soils for in situ life detection. Astrobiology 2017 Jul 13. doi: 10.1089/ast.2016.1535 PMID 28704064 OpenAccess

P3. Carr CE, Mojarro A, Hachey J, Saboda K, Tani J, Bhattaru SA, Smith A, Pontefract A, Zuber MT, Finney M, Doebler R, Brown M, Talbot R, Nguyen V, Bailey R, Ferguson T, Church G, Ruvkun G. Towards In Situ Sequencing for Life Detection. Aerospace Conference, 2017 IEEE. March 4-11, Big Sky, MT, USA, pp. 1-18. Session 2.07 In Situ Instruments for Landed Surface Exploration, Orbiters and Flybys. Paper # 2353 doi:10.1109/AERO.2017.7943896, Author Manuscript.

P2. Bhattaru SA, Tani J, Carr CE. Cold Testing of Conductive Bridging Random Access Memory for Space Applications. Aerospace Conference, 2017 IEEE. March 4-11, Big Sky, Montana. Session 7.09 Electronics for Extreme Environments. Paper # 2250. doi: 10.1109/AERO.2017.7943828, Author Manuscript.

P1. Carr CE, Mojarro A, Tani J, Bhattaru SA, Zuber MT, Doebler R, Brown M, Herrington K, Talbot R, Fuller C, Finney M, Church G, Ruvkun G. Advancing the Search for Extra-Terrestrial Genomes. Aerospace Conference, 2016 IEEE, March 5-12, Big Sky, Montana. doi: 10.1109/AERO.2016.7500859 

Selected Talks/Abstracts/Posters (27):

Carr CE. Resolving the History of Life on Earth by Seeking Life As We Know It on Mars. 2021 Exploration and Origins Colloquium, Georgia Tech (abstract and talk, presented 2021-02-18; distinguished speaker). Video: https://youtu.be/l3V7cFn9n6c

Christopher E. Carr, Daniel Duzdevich, Jack W. Szostak, Takahito Ohshiro, Gary Ruvkun, Jason M. Soderblom, and Maria T. Zuber. A Direct Search for Life As We Know It and Don’t Know It. 23rd SANKEN International Symposium, Awaji, Japan, January 9-10, 2020 (abstract, plenary talk) and 2020 ExplOrigins Colloquium, Georgia Institute of Technology, January 28, 2020 (abstract, plenary talk).

Carr CE. Towards In-Situ Detection of Informational Polymers for Life Detection on Ocean Worlds. Astrobiology Science Conference (2019). Invited Abstract + Talk.

Saboda KN, Rettberg P, Moeller R, Carr CE. Detecting Darwinian Evolution via Nanopore Sequencing. Astrobiology Science Conference, June 24-28, 2019. Abstract + Talk.

Carr CE, Bryan NB, Saboda K, Bhattaru SA, Ruvkun G, Zuber MT. Nanopore Sequencing at Mars, Europa, and Microgravity Conditions. Astrobiology Science Conference, June 24-28, 2019. Abstract + Poster.

Carr CE, Saboda K, Ruvkun G, Zuber MT, Wassmann M, Rettberg P, Moeller R. Informational polymers as unambiguous biomarkers for aqueous-based life. European Astrobiology Network Association (EANA), Berlin, Germany, 24-28 September, 2018 (abstract + talk).

Carr CE. The Limits of Life. Gary 66 Symposium, Boston, MA, June 11, 2018 (talk).

Carr CE, Ruvkun G, Zuber MT. Single Molecule Sensing and Sequencing for Life Detection Beyond Earth. Astrobiology Science Strategy for the Search for Life in the Universe, April 25-27, 2018, Washington DC (invited speaker and panelist).

Carr CE, Tani J, Ruvkun G, Zuber MT. Needs and Technologies to Enable the Direct Search for Life Beyond Earth. Technologies and Infrastructures Workshop for Planetary Exploration, Horizon 2061, EPFL, April 23-25, 2018 (talk and abstract).

Bryan N, Bhattaru SA, Saboda K, Ruvkun G, Zuber MT, Carr CE. DNA Sequencing During Simulated Mars, Europa, and Zero Gravity. MIT Media Lab Zero Gravity Flight Research Symposium. Jan 23, 2018 & MIT Media Lab Beyond the Cradle 2018, Mar 14, 2018 (poster).

Carr CE. Opportunities and Challenges for Life Detection via ‘Omics’ Approaches. ELSI/EON International Workshop on Life Detection, Earth Life Science Institute, Tokyo Institute of Technology, Japan. October 2017 (planned; invited speaker).

Carr CE. On the Search for Informational Polymers Beyond Earth. Earth Life Science Institute, Tokyo Institute of Technology, Japan. August 1, 2017 (invited seminar); also presented at Osaka University, Japan. July 20, 2017 (invited seminar).

Carr CE. Towards In Situ Sequencing for Life Detection. Future In-Space Operations Telecon, University of Texas, May 3, 2017 (http://spirit.as.utexas.edu/~fiso/telecon.htm; invited talk).

Carr CE, A. Mojarro, J. Hachey, A. Pontefract, R. Doebler, M. Brown, G. Ruvkun, and M. T. Zuber. Progress and Challenges for Life Detection via Nucleic Acid Sequencing. Astrobiology Science Conference, Mesa, Arizona, April 24–28, 2017. Abstract #3395 https://goo.gl/Tshurk

J. Hachey, A. Pontefract, M. T. Zuber, G. Ruvkun, C. E. Carr. Sequencing Nothing: Exploring Failure Modes of Nanopore Sensing and Implications for Life Detection. Astrobiology Science Conference, Mesa, Arizona, April 24–28, 2017. Abstract # 3454 https://goo.gl/AnEcJK

A. Mojarro, J. Hachey, R. Bailey, M. Brown, R. Doebler, G. Ruvkun, M. T. Zuber, C. E. Carr. Nucleic Acid Extraction and Sequencing from Low-Biomass Synthetic Mars Analog Soils. Lunar & Planetary Sci XLVIII, The Woodlands, Texas, March 21-25, 2017. Abstract # 1585 (poster) https://goo.gl/6xeAJJ

A. Pontefract, J. Hachey, A. Mojarro, V. K. Walker, H. Rowedder, T. F. Zhu, C. Lui, M. T. Zuber, G. Ruvkun, C. E. Carr. Understanding Habitability and Biosignature Preservation in a Hypersaline Mars Analog Environment: Lessons from Spotted Lake. Lunar & Planetary Sci XLVIII, The Woodlands, Texzas, March 21-25, 2017. Abstract # 1124 https://goo.gl/YBBRg6

Tani J, Ruvkun G, Zuber MT, Carr CE. On Neuromorphic Architectures for Efficient, Robust, and Adaptable Autonomy in Life Detection and Other Deep Space Missions. Planetary Science Vision 2050 Workshop, Washington, DC – Feb 27-Mar 1, 2017. Abstract # 8080, https://www.hou.usra.edu/meetings/V2050/pdf/8080.pdf, e-poster: https://goo.gl/1eWQsR

J. Hachey, A. Mojarro, J. Tani, A. Smith, A. Pontefract, S. A. Bhattaru, K. Saboda, G. Ruvkun, M. T. Zuber, and C. E. Carr. Single Molecule Sequencing for Life Detection Beyond Earth. Nanopore Community Meeting, New York, NY, December 1-2, 2016 (poster + talk), Video: https://nanoporetech.com/events/ncm16-lightning#195038663

Carr CE. On the Search for Informational Polymers Beyond Earth. Harvard Origins Initiative, Cambridge, MA, December 1, 2016 (invited chalk talk).

A. Mojarro, J. Hachey, J. Tani, A. Smith, S. A. Bhattaru, A. Pontefract, R. Doebler, M. Brown, G. Ruvkun, M. T. Zuber, C. E. Carr. SETG: Nucleic Acid Extraction and Sequencing for In Situ Life Detection on Mars. 3rd International Workshop on Instrumentation for Planetary Missions. Pasadena, CA, October 24-27, 2016. Abstract #4095. http://www.hou.usra.edu/meetings/ipm2016/pdf/4095.pdf

Carr CE, Hachey J, Smith A, Mojarro A, Pontefract A, Tani J, Zuber MT, Ruvkun G. Single molecule sequencing and the search for life beyond Earth. Cold Spring Harbor Asia, Microbiology and the Environment, Suzhou, China (September 26-30, 2016; invited speaker; session co-chair).

Mojarro, A., Ruvkun, G., Zuber, M. T., Carr, C. E. The Search for Extra-Terrestrial Genomes. Biosignatures and the Search for Life on Mars, Iceland, July 1-14, 2016 (poster).

Mojarro, A., Ruvkun, G., Zuber, M. T., Carr, C. E. The Search for Extra-Terrestrial Genomes: Nucleic Acid Extraction from Mars Analog Soils. 2016 Northeastern Geobiology Symposium, Poster.

Pontefract, A., Mojarro, A., Smith, A., Walker, V., Zuber, M., Ruvkun, G. and Carr, C.E. 2016. The Search for Unambiguous Biosignatures: DNA Extraction and Sequencing in Mars Analogue Environments. 2016 Northeastern Geobiology Symposium, Poster.

Mojarro A, Ruvkun G, Zuber MT, Carr CE. Methods for Extracting Nucleic Acids from Mars Analog Regolith. Lunar & Planetary Sci XLVII, The Woodlands, Texas, March 21-25, 2016. Abstract # 1643, https://www.hou.usra.edu/meetings/lpsc2016/pdf/1643.pdf

Mojarro, G. Ruvkun, M. T. Zuber, C. E. Carr. (2015) Human Exploration of Mars at Valles Marineris: The Past, Present, and Future of Life on Mars. First Landing Site/Exploration Zone Workshop for Human Missions to the Surface of Mars, Houston, TX, Oct 27-30. Abstract #1036, https://www.hou.usra.edu/meetings/explorationzone2015/pdf/1036.pdf

Addendum: Exploring Mars on Earth

To study the limits of life and to test instrumentation we have carried out fieldwork at terrestrial analogs of Mars. Examples include:

  • Volcán Copahue, Argentina, an active volcano with acidic iron-rich hydrothermal streams.
  • Haughton Impact Structure, Devon Island, Nunavut, Canada, a 20+ million year old impact crater in a polar desert with low mean annual temperatures.
  • Spotted Lake, Canada, a hypersaline magnesium sulfate rich analog for paleolakes on ancient Mars

Other significant contributions to date include: