1. Training Purposes Only JEM SPACE SEED 31407 JMU-081220 (Ver. 1.1) 1 Notice : This technical data is furnished on the condition that it will be used by and disclosed to the receiving Cooperating agency and its contractors and sub contractors only for the purposes of fulfilling the cooperating agency’s responsibilities under the Space Station Intergovernmental Agreement (IGA) and Memorandum of Understanding (MOU). It shall not be used for any other purpose, nor disclosed or retransferred to any other entity or government without prior written permission of the Japan Aerospace Exploration Agency (JAXA). Space Seed Experiment “Life Cycle of Higher Plants under Microgravity Conditions” JEM SPACE SEED 31407 Instructor:Takashi Yamasaki Toru Mori (yamasaki.takashi@jaxa.jp) (mori.tooru@jaxa.jp)

2. Training Purposes Only JEM SPACE SEED 31407 JMU-081220 (Ver. 1.1) 2 Objectives  Space Seed experiment overview  Space Seed experiment operations flow  Key points during Space Seed experiment operations  Space Seed experiment operations Explain the science objective of Space Seed experiment. List onboard outcomes from performing Space Seed experiment. Explain function of Plant Exp Unit. Explain function of Exp Laptop Terminal (ELT). Explain Space Seed operations flow regarding Plant Exp Unit. Explain function of KSC Fixation Tube (KFT). Explain what crew must check after attaching Plant Exp Unit on CBEF Turntable. Explain what crew must do in case fragments of plant tissue fly out of Glove Bag. Explain what crew must check before opening KFT Ziplock Bag. Explain what crew must take care while inserting plants in KFT. Explain what crew must take care while taking Plant Exp Sample Chamber out of Plant Exp Unit. Identify where crew must cut while harvesting plants. Explain what crew must not do while inserting plants in KFTs to prevent cross-contamination. Perform Space Seed experiment operations according to ODFs without instructor’s assistance. After this lesson the trainee will be able to:

3. Training Purposes Only JEM SPACE SEED 31407 JMU-081220 (Ver. 1.1) 3 Lesson Outline Contents: - Space Seed Experiment Overview - Space Seed Experiment Operations Flow - Key Points during Space Seed Experiment - Appendix Page: 4 8 10 11

4. Training Purposes Only JEM SPACE SEED 31407 JMU-081220 (Ver. 1.1) 4 Space Seed Experiment Overview  Plants grown in Space (Arabidopsis thaliana) - Plant Exp Sample Chamber (4) - Plant Exp Sample Chamber Bottom (4) - KFT (12) (Kennedy Space Center Fixation Tube) (RNAlater (4), Formaldehyde (2), Fromalin Acetic Acid Ethanol (2), Glutaraldehyde Paraformaldehyde (2), Glutaraldehyde Paraformaldehyde for Long Term (2))  Environment Gas - Plant Exp Gas Sampling Syringe (2)  Data - Temperature/Humidity Data - Video Images - Radiation Data - Microgravity Environment Data To study Life Cycle of Higher Plants under Microgravity Conditions. Space Seed experiment science objective Onboard Outcomes from Space Seed experiment

5. Training Purposes Only JEM SPACE SEED 31407 JMU-081220 (Ver. 1.1) 5 Space Seed Experiment Overview (cont’d) Plant Exp Unit - Plant Exp Unit is a kind of CBEF Canister to grow plants. - Plant Exp Unit has following components. - Plant Exp Sample Chamber - LEDs for growing plants - LEDs for lighting and a video camera - Water bag, Water Sensor and Water Supply Pump - Temperature/Humidity Sensor, Ventilation Pump, Heater and Circulation Fan - Plant Exp Sample Chamber also has following components. - Bio Dosimeter - Temperature Logger Bio Dosimeter Temperature Logger LED for growing plants Video Camera Water Bag Ventilation Pump Water Supply Pump Circulation Fan Temperature/Humidity Sensor Water Sensor Heater Plant Exp Chamber LED for lighting

6. Training Purposes Only JEM SPACE SEED 31407 JMU-081220 (Ver. 1.1) 6 Exp data files Plant Exp Unit Cell Biology Experiment Facility (CBEF) Installed in CBEF Incubator Units Ground Plant Exp Unit activation/deactivation Commands Plant Exp Unit on/off status, Temp of circuit board Exp Laptop Terminal (ELT) (Think Pad A31p) ELT Data Cable Temperature/Humidity of Sample Chamber, Pump/Fan status, etc. Communication Check Experiment start/stop, etc. Parameter files Plant Exp Units in CBEF Micro-G Incubator Unit Plant Exp Units in CBEF 1G Incubator Unit Exp Laptop Terminal Space Seed Experiment Overview (cont’d) - Exp Laptop Terminal is a laptop computer for controlling Plant Exp Unit in CBEF.

7. Training Purposes Only JEM SPACE SEED 31407 JMU-081220 (Ver. 1.1) 7 Intentionally Blank

8. Training Purposes Only JEM SPACE SEED 31407 JMU-081220 (Ver. 1.1) 8 CBEF Plant Exp Unit :Crew operations :Ground operations LEGEND: CBEF Activation/Environment Control Plant Exp Unit Activation Experiment Start by ELT Plant Exp Unit Installation (8) -Microgravity Environment Data Exp Laptop Terminal Measurement Start Space Seed Experiment Operations Flow Dehumidifier InstallationELT Setup/Activation MMA Experiment Stop by ELT Plant Exp Unit Deactivation Plant Exp Unit Removal (4) Harvest (Short/Long Term) CBEF Deactivation Dehumidifier Removal ELT Deactivation/Removal Measurement Stop Repeat for Long Term Experiment MLT Activation MLT Deactivation MELFI Insertion Tasks just before or during Long Term Experiment Dehumidifier Exchange Water Supply/Gas Sampling ELT Check (periodically) -Temperature/Humidity Data -Video Image -Radiation Data -Cultivation (Short Term: approx. 30 days) (Long Term: approx. 60 days) Exp Laptop Terminal Plant Exp Unit After Long Term Experiment Dehumidifier Gas Sampling Syringe PEU Program Installation Before Short Term experiment

9. Training Purposes Only JEM SPACE SEED 31407 JMU-081220 (Ver. 1.1) 9 Space Seed Experiment Operations Flow (Cont’d) Harvest Take out Sample Chambers from Plant Exp Units Put Sample Chambers, KFTs, and Harvest Kit in Glove Bag Open Sample Chamber Harvest plants Put plants in KFTs Fix plants by KFTs Take out Sample Chambers, KFTs, and Harvest Kit from Glove Bag Glove Bag Space Seed Harvest Kit KSC Fixation Tube Fix plants Put plants in KFT Harvest plants Open Sample Chamber Take out Sample Chamber

10. Training Purposes Only JEM SPACE SEED 31407 JMU-081220 (Ver. 1.1) 10 Key Points during Space Seed Experiment NOTE: Underlined key points have been covered by CBEF Advanced Training. CAUTION: Payload Safety Review Panel has not been completed yet. CategoryEquipmentKey Points Crew Safety Electrical ShockSample Tray, Centrifuge Before attaching/detaching the Plant Exp Units to/from the CBEF Sample Tray/Centrifuge, power to Plant Exp Units must be turned off by switches on the Sample Tray/Centrifuge. Entanglement with rotating device Incubator Unit 1G Verify visually through the Observation Window that CBEF turntable does not rotate before opening the door of the CBEF Incubator Unit 1G. CentrifugeWhen attaching/detaching the Plant Exp Units to/from the CBEF 1G Centrifuge, lock turntable of the Centrifuge by stopper pin. Release of fragments from rotating device Centrifuge, Plant Exp Unit Verify no gap exists between base plate of Plant Exp Unit and CBEF Turntable. Biomaterial Leakage to Cabin Glove BagVacuum fragments of plant tissues in case they fly out of Glove Bag. Release of hazardous chemicals KSC Fixation Tube- Check KFT seals (2 sets of triple O-ring seals on KFT Base Plug and KFT Expansion Plug) have good compression against Main Tube and there are no leaks before opening KFT Ziplock Bag. - Insert plants into KFT so that they are no longer than the Sample Tube and KFT Actuator O-rings will NOT catch them. Contamination due to hazardous material leakage to Cabin CBEF DehumidifierVerify that there is no leakage before taking out CBEF Dehumidifier from Ziplock Bag. Time Critical OperationN/A Dos and Don’tsIncubator UnitDo NOT touch inside of CBEF Incubator Units with bare hand to keep them clean. Sample Tray, Centrifuge Be careful not to pinch fingers when releasing drawer locks of slide rails on CBEF Sample Tray/Centrifuge. Plant Exp UnitDo NOT pull Plant Exp Sample Chamber all the way out of Plant Exp Unit. Or cable/hoses may be snapped. N/A (Plant Sample) - Cut under leaves while harvest. - Do NOT insert unidentifiable plants in KFTs to prevent cross-contamination. Difficulty in OpsN/A Possible Mis-OpsN/A

11. Training Purposes Only JEM SPACE SEED 31407 JMU-081220 (Ver. 1.1) 11 Appendix - Harvest for Short Term Experiment - Harvest for Long Term Experiment - Harvest in case of KFT leakage - Acronyms

12. Training Purposes Only JEM SPACE SEED 31407 JMU-081220 (Ver. 1.1) 12 MELFI -95 deg C Harvest for Short Term Experiment Sample Chamber Top (4) Trash A side B side A side B side Half of B side A side Half of B side A side Half of B side Sample Chamber Bottom (4) KFT RNAlater (4) KFT Formaldehyde (2) KFT Formalin Acetic Acid Ethanol (2) KFT Glutaraldehyde Paraformaldehyde (2) MELFI 2 deg C CBEF Micro-G Incubator Unit CBEF 1G Incubator Unit KFT RNAlater (4) Relocation 5 days later Harvest for Short Term experiment (approx. 30 days) Remain in CBEF Incubator Unit for Long Term Experiment S/N 001 S/N 003 S/N 005 S/N 007

13. Training Purposes Only JEM SPACE SEED 31407 JMU-081220 (Ver. 1.1) 13 Harvest for Long Term Experiment Harvest for Long Term experiment (approx. 60 days) CBEF Micro-G Incubator Unit CBEF 1G Incubator Unit Sample Chamber (4) Stowed in MELFI from Short Term Experiment MELFI -95 deg C Sample Chamber Bottom (4) KFT Formaldehyde (2) KFT Formalin Acetic Acid Ethanol (2) KFT Glutaraldehyde Paraformaldehyde (2) MELFI 2 deg C KFT RNAlater (4) Half of B side Sample Chamber Bottom (2) Sample Chamber Top (2) KFT Glutaraldehyde Paraformaldehyde for Long Term (2) S/N 004 S/N 008 S/N 004 S/N 008 S/N 002 S/N 006

14. Training Purposes Only JEM SPACE SEED 31407 JMU-081220 (Ver. 1.1) 14 MELFI -95 deg C KFT RNAlater (3) MELFI 2 deg C CBEF Micro-G Incubator Unit CBEF 1G Incubator Unit KFT RNAlater (3) Relocation 5 days later Harvest in case of KFT RNALater leakage Remain in CBEF Incubator Unit for Long Term Experiment Harvest in case of KFT leakage - There are 5 KFT RNALater (4 used, 1 spare). - If 1 KFT RNALater has leakage, use spare KFT RNALater. -If 2 KFT RNALater have leakage, plant samples from 2 Plant Exp Sample Chamber A side (S/N 005, 007) will be fixed in 1 KFT RNA Later. A side S/N 001 S/N 003 S/N 005 S/N 007

15. Training Purposes Only JEM SPACE SEED 31407 JMU-081220 (Ver. 1.1) 15 MELFI -95 deg C B side KFT Formaldehyde (2) MELFI 2 deg C CBEF Micro-G Incubator Unit CBEF 1G Incubator Unit Remain in CBEF Incubator Unit for Long Term Experiment Harvest in case of KFT leakage (Cont’d) Harvest in case of KFT Formaldehyde leakage - There are 3 KFT Formaldehyde (2 used, 1 spare). - If 1 KFT Formaldehyde has leakage, use spare KFT Formaldehyde. S/N 001 S/N 005

16. Training Purposes Only JEM SPACE SEED 31407 JMU-081220 (Ver. 1.1) 16 - There are 2 KFT Formalin Acetic Acid Ethanol (2 used). - If 1 KFT Formalin Acetic Acid Ethanol has leakage, plant sample from B side of Plant Exp Sample Chamber S/N 007 will be fixed by KFT Glutaraldehyde Paraformaldehyde. MELFI -95 deg C Half of B side B side KFT Formalin Acetic Acid Ethanol (1) KFT Glutaraldehyde Paraformaldehyde (2) MELFI 2 deg C CBEF Micro-G Incubator Unit CBEF 1G Incubator Unit Remain in CBEF Incubator Unit for Long Term Experiment Harvest in case of KFT leakage (Cont’d) Harvest in case of KFT Formalin Acetic Acid Ethanol leakage S/N 003 S/N 007

17. Training Purposes Only JEM SPACE SEED 31407 JMU-081220 (Ver. 1.1) 17 MELFI -95 deg C Half of B side B side KFT Formalin Acetic Acid Ethanol (2) KFT Glutaraldehyde Paraformaldehyde (1) MELFI 2 deg C CBEF Micro-G Incubator Unit CBEF 1G Incubator Unit Remain in CBEF Incubator Unit for Long Term Experiment Harvest in case of KFT leakage (Cont’d) Harvest in case of KFT Glutaraldehyde Paraformaldehyde leakage S/N 003 S/N 007 - There are 2 KFT Glutaraldehyde Paraformaldehyde (2 used). - If 1 KFT Glutaraldehyde Paraformaldehyde has leakage, plant sample from B side of Plant Exp Sample Chamber S/N 007 will be fixed by KFT Formalin Acetic Acid Ethanol.

18. Training Purposes Only JEM SPACE SEED 31407 JMU-081220 (Ver. 1.1) 18 CBEF Micro-G Incubator Unit CBEF 1G Incubator Unit Sample Chamber (4) MELFI -95 deg C KFT Formaldehyde (2) MELFI 2 deg C Half of B side Sample Chamber Bottom (1) Sample Chamber Top (2) KFT Glutaraldehyde Paraformaldehyde for Long Term (1) S/N 004 Harvest in case of KFT leakage (Cont’d) Harvest in case of KFT Glutaraldehyde Paraformaldehyde for long term leakage - There are 2 KFT Glutaraldehyde Paraformaldehyde for long term (2 used). - If 1 KFT Glutaraldehyde Paraformaldehyde for long term has leakage, plant sample from half of B side of Plant Exp Sample Chamber S/N 008 will be fixed by KFT Formaldehyde spare for short term experiment. S/N 004 S/N 008 S/N 002 S/N 006 KFT Formaldehyde (1) (Spare for short term) Half of B side Sample Chamber Bottom (1) S/N 008 Stowed in MELFI from Short Term Experiment

19. Training Purposes Only JEM SPACE SEED 31407 JMU-081220 (Ver. 1.1) 19 Acronyms CBEF ELT KSC KFT LED MMA MLT MELFI ODF Cell Biology Experiment Facility Experiment Laptop Terminal Kennedy Space Center KSC Fixation Tube Light Emitting Diode Microgravity Measurement Apparatus MMA Laptop Terminal Minus Eighty degree Laboratory Freezer for ISS Operations Data File