Towards in situ sequencing for life detection

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.