S. cerevisiae BY4743 grown in YPD in HARV bioreactor modeling low-shear microgravity
Biomaterial provider
Invitrogen
Treatment protocol
None
Growth protocol
T=30C; grown for 5 generations; harvested in log phase in YPD
Extracted molecule
total RNA
Extraction protocol
Cells were collected from the HARVs, the OD600 was measured, and the cells were pelleted by centrifugation at 5000 X g for 5 min at room temperature. The cell pellets were flash frozen in dry ice/ethanol and stored at -80 oC. Total RNA was extracted using the QIAGEN® RNeasy® Midi Kit (QIAGEN, Inc., Valencia, CA) according to the manufacturer’s instructions with the following modification to the cell lysis step. We added 0.1mm diameter sterile zirconia/silica beads (BioSpec Products, Inc., Bartlettsville, OK) to the yeast cells in buffer RLT, then bead beat in the Savant Fast Prep bead beater (Savant Instruments, Inc., Farmingdale, NY) at speed 6.5 for 45 seconds, and immediately cooled the samples on ice. Samples for microarray analysis were further cleaned-up and concentrated using the QIAGEN® RNeasy® MinElute Cleanup Kit (QIAGEN, Inc., Valencia, CA). RNA quality was evaluated by 1.2% formaldehyde/agarose gel electrophoresis according to the QIAGEN protocol in Appendix H of the RNeasy® Mini Handbook.
Label
biotin
Label protocol
Approximately 6 µg of total RNA was processed to produce biotinylated cRNA targets using Affymetrix One-Cycle Target prep protocol.
Hybridization protocol
standard Affymetrix procedures
Scan protocol
standard Affymetrix procedures
Description
This sample is grown in low-shear modeled microgravity (LSMMG).
