|
| Status |
Public on Dec 01, 2018 |
| Title |
FLIGHT BRIC A PDFU-2 302 WT |
| Sample type |
RNA |
| |
|
| Source name |
wildtype_space_flown_solid media plates; BRIC-17; Dragon,ISS; BRIC-A PDFU-2
|
| Organism |
Arabidopsis thaliana |
| Characteristics |
flight status: space flown
cultivar: Col-0
genotype: WT
material type: Tissue culture cells
tissue source: derived from hypocotyls of WT plant
age: 14 days old
biosource type: RNAlater in-flight fixed_sample
|
| Treatment protocol |
The BRIC-17 experiment was turned over to payload engineers in the SLSL at KSC 48 hours before the scheduled launch time. The payload launched on Dragon cargo spacecraft, SpaceX CRS-2 on March 1st, 2013, at 10:10 EST, and achieved orbit approximately twelve minutes later. Dragon berthed to the ISS on the March 3rd, 2013. The biology was in orbit 10 days before being fixed on orbit with RNAlater. The Dragon unmanned spacecraft unberthed the ISS and splashed into Pacific on the March 26th and the biology was returned to the Kennedy Space Center and handed to the PIs at SLSL on the April the 2nd.
|
| Growth protocol |
General biology operations Two Arabidopsis thaliana Col-0 cell culture lines were used: Wild Type (WT) and ARG1 KO SALK_024542C line (ARG1 T-DNA insertion knockout), all cell cultures derived from hypocotyls. Tissue culture cells were plated on 60mm round, nutrient agar Petri plates cultured in a horizontal position until turnover and installation into the BRIC hardware. All of the sub-culturing on the plates was conducted in the laboratory of the PIs and the University of Florida. Plates were planted in a laminar flow hood and stored in sterile Biotransfer Boxes (BT). Plates with cells were kept in the dark and at ambient temperature. Preparation of specialized plates and biology Petri plates. Because of their configuration within the BRIC flight hardware, sterility was maintained on all surfaces (interior and exterior) of the Petri plates. The media in the plates for BRIC had to be 5mm deep or less to accommodate the biology and the PDFU cover hardware. For a 60mm dish, this calculates out to 6.7 mLs. After pouring, the plates are allowed to solidify, and then transferred to a sterile Biotransporter (to maintain sterility of external surface) until planting. Preparation of cell culture plates. The liquid media from a sterile cell suspension was decanted, and the cells were washed once with fresh liquid media, and then decanted again. A sterile scoop was used to place about 1 gram of cells in the surface on the plate and disperse evenly across the surface. The cover was replaced, but not taped, and transferred to a sterile BT. The BT was wrapped in black cloth and stored at room temperature. The same day plates were transported by car to the Kennedy Space Center, NASA for the same night (3:00 AM) Turn Over. The ground control plates were turned over 48 hours later. BRIC Spaceflight Hardware The hardware for this experiment was the NASA BRIC-LED (Biological Research In a Canister – Light Emitting Diode). The LED component of the hardware was disabled and all biology was held in the dark for the duration of the experiment. The BRIC-LED canisters hold six 60 mm diameter Petri plates in small sub-compartments called PDFUs (Petri Dish Fixation Units). The PDFUs allow an externally activated fixation step without manipulating the samples. We were allocated 10 PDFUs: 5 in BRIC A and 5 in BRIC B. The 2 replicate plates of WT cultured cells and the 3 replicate plates of ARG1 KO were distributed among the BRIC A canister. The 2 replicate plates of WT cultured cells were distributed among the BRIC B canister. One of the PDFU slots in each BRIC was used to hold a HOBO™ data logger to record temperature. The HOBO temperature data collection begins immediately after the biology is sealed in the BRICs, 24 hours before launch. An additional set of Petri plates within BRIC hardware, identical to that launched for Spaceflight, was prepared for the Ground Control. The Ground Control was housed in the OES (Orbital Environmental Simulator) chamber in the Spaceflight Life Sciences Laboratory (SLSL) at Kennedy Space Center (KSC). The Ground Control was initiated with a precise 24 hour delay to enable the OES environment to be programmed with the environmental profile taken from telemetry of the Space Shuttle. Nominally, the Ground Control BRIC hardware and samples experienced the same external environment profile as the hardware and samples on the Space Shuttle. Environmental parameters that are controlled on the OES and made identical to flight are cabin temperature, humidity and CO2 concentration.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total cellular RNA was extracted using RNeasy Mini Kit (QIAGEN)
|
| Label |
biotin
|
| Label protocol |
cDNA was synthesized using Ovation Pico WTA System (NuGEN Technologies Inc.) and cDNA was labeled using Encore Biotin Module (NuGEN Technologies Inc.)
|
| |
|
| Hybridization protocol |
Amplified and labeled cDNA (5 µg/sample) was fragmented and hybridized with rotation onto Affymetrix GeneChip Arabidopsis ATH1 Genome Arrays for 16h at 45°C. Arrays were washed on a Fluidics Station 450 (Affymetrix) with the Hybridization Wash and Stain Kit (Affymetrix) and the Washing Procedure FS450_0004.
|
| Scan protocol |
scan was performed using Affymetrix GeneChip Scanner 3000 7G
|
| Description |
FA2
|
| Data processing |
The MAS5 data was generated by using the Affymetrix Expression Console software using default analysis settings and global scaling as normalization method.
|
| |
|
| Submission date |
May 13, 2016 |
| Last update date |
Dec 01, 2018 |
| Contact name |
Robert J. Ferl |
| E-mail(s) |
ferllabuf@gmail.com
|
| Phone |
352-273-8030
|
| Organization name |
University of Florida
|
| Department |
Horticultural Sciences
|
| Lab |
Ferl's lab
|
| Street address |
1301 Fifield Hall PO Box 110690
|
| City |
Gainesville |
| State/province |
Florida |
| ZIP/Postal code |
32611 |
| Country |
USA |
| |
|
| Platform ID |
GPL198 |
| Series (2) |
| GSE81442 |
Arg1 functions in the physiological adaptation of undifferentiated plant cells to spaceflight |
| GSE95388 |
HSFA2 functions in the physiological adaptation of undifferentiated plant cells to spaceflight microgravity environment. |
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