Three years after the experiment was conceived and designed by high school students, astronauts have demonstrated the first successful use of CRISPR/Cas9 genome editing in a microgravity environment aboard the International Space Station (ISS).
In 2018, Minnesota high school students David Li, Michelle Sung, Aarthi Vijayakumar and Rebecca Li won the Genes in Space competition, an annual challenge developed by Boeing and miniPCR bio that seeks to inspire students in grades 7 through 12 to design DNA experiments that address a challenge in space exploration. Their experiment focused on developing a workflow to study DNA damage repair in microgravity.
Now, ISS astronauts Christina Koch and Nick Hague, along with Earth colleagues (and controls) NASA’s Sarah Castro-Wallace and Sarah Stahl-Rommel, have just published the successful results in PLOS One.
"Over the past 3 years, we've worked and watched as an idea we discussed at midnight one day became a fully fledged experiment conducted on the ISS, and now finally shared with the scientific community," said a now-20-year-old Vijayakumar. "This project and process has taught us so much."
Astronauts traveling outside Earth's protective atmosphere face increased risk of DNA damage due to the ionizing radiation that permeates space. The most significant damage—double-strand breaks, which occur when the phosphate backbones of both DNA strands are hydrolyzed—increase the risk of cancer and other deadly diseases.
And while scientists have a workflow to repair DNA damage on Earth, the unique conditions of microgravity prevented such a process in space—until now.
The first challenge the team faced was transforming the yeast cells (Saccharomyces cerevisiae) onboard the ISS, which required a smaller volume of culture to be dispensed on an agar plate via micropipette compared with the traditional Earth/ground method.
“By transferring small volumes, the surface tension was sufficient to keep the yeast suspension attached to the agar before being spread by the force of a sterile plastic spreader,” the authors explain in their paper.” Ground controls at NASA were prepared in parallel using the same, non-traditional method.
Six days later, the ISS crew reported 4 red colonies and 6 white colonies, while the ground controls contained 8 red and 29 white colonies. According to the study, the red phenotype is indicative of successful CRISPR/Cas9 mutagenesis.
Subsequent PCR and DNA sequencing confirmed that the 4 white colonies grown during spaceflight and on the ground yielded 1.3 million and 1 million total reads, respectively. Meanwhile, sequencing of the 4 red colonies yielded more than 5 million (flight) and 2 million (ground) total reads.
While these numbers are lower than what is typically seen in traditional S. cerevisiae transformation experiments, the efficiency was still sufficient to enable the detection of CRISPR edited cell—thus confirming the first CRISPR/Cas9 genome editing event in space.
The authors attribute this decline in efficiency to the fact that the new microgravity-friendly porotocal relied on reduced sample volume and reagents that were premixed and frozen, rather than prepared fresh as is typical.
The scientists said they hope the technique will now enable extensive research into DNA repair in space.
"Future studies could focus on improving the efficiency of the transformation protocol described here to allow a more extensive investigation of the frequency of homologous recombination (HR) compared to non-homologous end joining (NHEJ) DNA repair in microgravity conditions,” they write.
But that's not all.
The technique can serve as the foundation for investigations into numerous other molecular biology topics related to long-term space exposure and exploration, which is especially important as the commercial space race heats up and humans envision a future with Mars as a home planet.
Photo: NASA astronaut Christina Kock performing the experimental procedure aboard the International Space Station. Credit: Sebastian Kraves