Genes in Space-6 (Genes in Space-6) - 01.30.19

Overview | Description | Applications | Operations | Results | Publications | Imagery

ISS Science for Everyone

Science Objectives for Everyone
Deoxyribonucleic acid (DNA) damage caused by increased exposure to radiation can affect the long-term health of astronauts. Genes in Space-6 determines the optimal DNA repair mechanisms that cells use in the spaceflight environment. The investigation evaluates the entire process in space for the first time by inducing DNA damage in cells and assessing mutation and repair at the molecular level using the miniPCR and the Biomolecule Sequencer tools aboard the space station.
Science Results for Everyone
Information Pending

The following content was provided by Sarah Wallace, Ph.D., and is maintained in a database by the ISS Program Science Office.
Experiment Details

OpNom: Genes in Space-6

Principal Investigator(s)
David Li, Amplyus, Cambridge, MA, United States
Michelle Sung, Amplyus, Cambridge, MA, United States
Aarthi Vijayakumar, Amplyus, Cambridge, MA, United States
Rebecca Li, Amplyus, Cambridge, MA, United States

Co-Investigator(s)/Collaborator(s)
David SCOTT Copeland, The Boeing Company, Pasadena, TX, United States
Sarah Wallace, Ph.D., NASA Johnson Space Center, Houston, TX, United States
Sarah Stahl, M.S., NASA Johnson Space Center, Houston, TX, United States
Emily Gleason, Ph.D., Amplyus, Cambridge, MA, United States
Ezequiel Alvarez Saavedra, Ph.D., Amplyus, Cambridge, MA, United States
Sebastian Kraves, Ph.D., Amplyus, Cambridge, MA, United States
Deniz Atabay, Massachusetts Institute of Technology, Cambridge, MA, United States
Nicole Nichols, Ph.D., Amplyus, Ipswich, MA, United States
Ashley Luck, Ph.D., Amplyus, Ipswich, MA, United States

Developer(s)
Boeing, Houston, TX, United States

Sponsoring Space Agency
National Aeronautics and Space Administration (NASA)

Sponsoring Organization
National Laboratory Education (NLE)

Research Benefits
Information Pending

ISS Expedition Duration
March 2019 - October 2019

Expeditions Assigned
59/60

Previous Missions
Genes in Space -1, -2, -3, -4, -5

^ back to top

Experiment Description

Research Overview

  • Genes in Space-6 assesses the yeast, Saccharomyces cerevisiae, ability to repair deoxyribonucleic acid (DNA) double strand breaks induced by clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 in a space environment.
  • Radiation in the space environment may cause breaks in deoxyribonucleic acid (DNA), known as double strand breaks. Cells repair these breaks but are prone to errors, causing insertions or deletions (indels) of DNA bases. Indel buildup may have detrimental effects, such as cancer.
  • Unlike previous investigations, Genes in Space-6 causes double strand breaks in a space environment with CRISPR/Cas9, so all repairs, which happen almost immediately after the breaks, are done in the space environment.
  • For the first objective, ground-based, edited genomic DNA is launched to the International Space Station (ISS). Flight-based experimentation only involves amplification and sequencing of the edited DNA section.
  • For the second objective, mutant and wild type S. cerevisiae strains are launched. The mutant is characterized to preferentially employ NHEJ, while the wild type (WT) employs HR. Both strains are set up in form of stasis and new populations are cultured aboard the ISS. The CRISPR/Cas-9 system is activated and the guide ribonucleic acid (gRNAs) targets a sequence within the ADE2 gene. Following time to allow for repair, DNA is extracted from both strains using miniPCR. Following a bead-based cleanup of the extracted DNA, a DNA region spanning the ADE2 gene is amplified using miniPCR. Another bead-based cleanup, as well as DNA library preparation, is completed prior to the final step of DNA sequencing.

Description

Genes in Space-6 studies the ability of yeast to repair induced double strand breaks in a space environment.
 
Spaceflight radiation may result in deoxyribonucleic acid (DNA) breaks, referred to as double strand breaks. Cells have many different ways to repair these breaks, but these repair processes are error prone and give rise to insertions and deletions (indels) of DNA bases (the A, G, T, and C). The long-term buildup of these indels may result in the development of cancer and other degenerative disease. Furthermore, the genes that are turned on and off in response to spaceflight (changes in gene expression) may affect the repair process and the accumulation of indels. Since astronauts may be exposed to increased radiation on exploration class missions, it is therefore important to understand 1) how the change in gene expression in response to spaceflight impacts cellular DNA repair processes and 2) what is the least error prone mechanism of repair in space.
 
Previous investigations have aimed to understand DNA repair in space. For this, breaks have been caused on Earth and the cells launched frozen for culture and repair once in the spaceflight environment. There are issues with this line of investigation because repair is initiated almost instantaneously: by the time the cells have been frozen, some repair has likely already occurred. This work is novel in that the entire process, damage through repair, is evaluated in space. To accomplish this goal, the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system is used to cause controlled double strand breaks. This system provides the ability to cause breaks in the DNA at exact locations, which allows for a simpler approach to determining repair efficiency and the presence of indels. The molecular tools on board, miniPCR, which can perfrom a polymerase chain reaction and the Biomolecule Sequencer, may then be used to measure indels within the DNA.
 
This investigation’s first objective is the Indel Sequencing Proof of Concept. A simple-to-perform method is used as a proof-of-concept to collect baseline data toward indel sequencing in space that then serves as a means of control for the cell-based experiments. To execute objective 1, DNA is extracted from cells that have experienced CRISPR/Cas9 induced double strand breaks and the corresponding repair on Earth. Since the location of the DNA damage - and thus indels - is known, miniPCR amplifies this section of the DNA. The amplified DNA then is prepared for sequencing, and indels are identified using the Biomolecule Sequencer.
 
The investigation’s second objective is called DNA Repair and Indel Generation in Response to Spaceflight. Two strains of Saccharomyces cerevisiae (commonly referred to as baker’s yeast) that preferentially use two different means of double strand break repair are sent to the International Space Station (ISS). To execute this objective, the crew performs the following: 1) Culture S. cerevisiae through incubation at ambient ISS temperature. 2) Activate the CRISPR/Cas-9 system to generate double strand breaks. 3) Allow repairs to occur through incubation at ambient ISS temperature. 4) Extract DNA using miniPCR. 5) Amplify the repaired section of DNA with miniPCR. 6) Prepare the amplified DNA for sequencing. 7) Sequence Indels using the Biomolecule Sequencer.

^ back to top

Applications

Space Applications
DNA damage caused by space radiation has been linked to increased risk of cancer and other degenerative diseases. Increased understanding of how cells repair this DNA damage in microgravity can contribute to procedures to protect astronauts on the space station and to future missions beyond low-Earth orbit.

Earth Applications
The simple, effective procedures needed for a remote, constrained resource environment such as the space station may have applications for protecting people from radiation and other hazards in remote and harsh locations on Earth.

^ back to top

Operations

Operational Requirements and Protocols

Objective 1: Deoxyribonucleic acid (DNA) extracted from cells that have experienced double strand breaks induced by clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 and the corresponding repair on Earth are retrieved from Minus Eighty-Degree Laboratory Freezer for ISS (MELFI) and undergo amplification using miniPCR to perfrom a polymerase chain reaction. Following amplification, the DNA is prepped (via pipetting and magnetic bead use) and sequenced using the Biomolecule Sequencer.
 
Objective 2: Saccharomyces cerevisiae cells are activated to grow either through transfer to a new tube (via pipetting) or by thawing from a frozen state. The cells grow during incubation at ambient International Space Station (ISS) temperature. CRISPR/Cas-9 is activated (via pipetting) within the cells. The cells are allowed to incubate for an additional amount of time. DNA is extracted with miniPCR and cleaned up (via pipetting and magnetic bead use) for amplification again with the use of miniPCR. Following amplification, the DNA is prepped (via pipetting and magnetic bead use) and sequenced using the Biomolecule Sequencer.

^ back to top

Decadal Survey Recommendations

Information Pending

^ back to top

Results/More Information

Information Pending

^ back to top

Related Websites
Genes in Space
miniPCR

^ back to top


Imagery