The increasing availability of flights on suborbital rockets creates new avenues for the study of spaceflight effects on biological systems, particularly of the transitions between hypergravity and microgravity. This paper presents an initial comparison of the responses of Arabidopsis thaliana to suborbital and atmospheric parabolic flights as an important step toward characterizing these emerging suborbital platforms and their effects on biology. Transcriptomic profiling of the response of the Arabidopsis ecotype Wassilewskija (WS) to the aggregate suborbital spaceflight experiences in Blue Origin New Shepard and Virgin Galactic SpaceShipTwo revealed that the transcriptomic load induced by flight differed between the two flights, yet was biologically related to traditional parabolic flight responses. The sku5 skewing mutant and 14-3-3κ:GFP regulatory protein overexpression lines, flown in the Blue Origin and parabolic flights, respectively, each showed altered intra-platform responses compared to WS. An additional parabolic flight using the F-104 Starfighter showed that the response of 14-3-3κ:GFP to flight was modulated in a similar manner to the WS line. Despite the differing genotypes, experimental workflows, flight profiles, and platforms, differential gene expression linked to remodeling of central metabolic processes was commonly observed in the flight responses. However, the timing and directionality of differentially expressed genes involved in the conserved processes differed among the platforms. The processes included carbon and nitrogen metabolism, branched-chain amino acid degradation, and hypoxic responses. The data presented herein highlight the potential for various suborbital platforms to contribute insights into biological responses to spaceflight, and further suggest that in-flight fixation during suborbital experiments will enhance insights into responses during each phase of flight.

Suborbital spaceflights now enable human-tended research investigating short-term gravitational effects in biological systems, eliminating the need for complex automation. Here, we discuss a method utilizing KSC Fixation Tubes (KFTs) to both carry biology to suborbital space as well as fix that biology at certain stages of flight. Plants on support media were inserted into the sample side of KFTs preloaded with RNAlater in the fixation chamber. The KFTs were activated at various stages of a simulated flight to fix the plants. RNA-seq analysis conducted on tissue samples housed in KFTs, showed that plants behaved consistently in KFTs when compared to petri-plates. Over the time course, roots adjusted to hypoxia and leaves adjusted to changes in photosynthesis. These responses were due in part to the environment imposed by the encased triple containment of the KFTs, which is a requirement for flight in human spacecraft. While plants exhibited expected reproducible transcriptomic alteration over time in the KFTs, responses to clinorotation during the simulated flight suggest that transcriptomic responses to suborbital spaceflight can be examined using this approach.

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The Cosmic Ray Exposure Sequencing Science (CRESS) payload system was a proof of concept experiment to assess the genomic impact of space radiation on seeds. CRESS was designed as a secondary payload for the December 2016 high-altitude, long-duration south polar balloon flight carrying the Boron and Carbon Cosmic Rays in the Upper Stratosphere (BACCUS) experiment. Investigation of the biological effects of Galactic Cosmic Radiation (GCR), particularly those of ions with High-Z and Energy (HZE), was of interest due to the genomic damage this type of radiation inflicts. The biological effects of radiation above Antarctica (ANT) were studied using Arabidopsis thaliana seeds and compared to a simulation of GCR at Brookhaven National Laboratory (BNL) and to laboratory control seeds. The CRESS payload was broadly designed to 1U CubeSat specifications (10 cm × 10 cm × 10 cm, ≤1.33 kg), maintained 1 atm internal pressure, and carried an internal cargo of 580,000 seeds and twelve CR-39 Solid-State Nuclear Track Detectors (SSNTDs). Exposed BNL and ANT M0 seeds showed significantly reduced germination rates and elevated somatic mutation rates when compared to non-irradiated controls, with the BNL mutation rate also being higher than that of ANT. Genomic DNA from plants presenting distinct aberrant phenotypes was evaluated with whole-genome sequencing using PacBio SMRT technology, which revealed an array of structural genome variants in the M0 and M1 plants. This study was the first whole-genome characterization of space-irradiated seeds and demonstrated both the efficiency and efficacy of Antarctic long-duration balloons for the study of space radiation effects on eukaryote genomes.

Biological experiments on-orbit that demonstrate the effects of gravity on plants require precise control of the initiation of plant development. Preserving seed dormancy is critical to experiments that endeavor to study the effects of the orbital environment, independent of contributions from either a normal gravity, or launch. However, spaceflight experiments are often tightly constrained with respect to the configuration of the biology and associated hardware, and it is rarely possible to launch dry seeds separated from their growth substrate. Described here are techniques established to maintain viable seeds that can remain dormant for up to a month at room temperature, and hydrated on the surface of solid, Phytagel growth medium. The configuration can also accommodate a brief (less than one minute) exposure to light during the quiescent period for quick inspection for any breaks in dormancy, and for contamination. The data presented outline the preparation of sealed, Phytagel media plates of dormant Arabidopsis thaliana seed that can be activated in situ when unwrapped and installed within a lighted growth habitat. These protocols were developed primarily for spaceflight scenarios where seeded plates must be prepared ahead of time and kept at ambient temperatures. However, these protocols can be adapted for any field application where it is desirable to transport dormant, seeded plates to a remote location where it would not be possible to prepare sterile culture plates.