Isolated RNA from a subset of Ronit’s Crassostrea gigas ctenidia samples (see Ronit’s notebook for experiment deets):
- D01 C
- D02 C
- D19 C
- D20 C
- T01 C
- T02 C
- T19 C
- T20 C
RNA was isolated using RNAzol RT (Molecular Research Center) in the following way:
- Unweighed, frozen tissue transferred to 500uL of RNAzol RT and immediately homogenized with disposable pestle.
- Added additional 500uL of RNAzol RT and vortexed to mix.
- Added 400uL of 0.1% DEPC-treated H2O, vortexed and incubated 15mins at RT.
- Centrifuged 12,000g for 15mins at RT.
- Transferred 750uL of supernatant to clean tube (discarded remainder), added 1 volume (750uL) of isopropanol, vortexed, and incubated at RT for 10mins.
- Centrifuged 12,000g for 10mins at RT.
- Discarded supernatant.
- Washed pellet with 75% ethanol (made with 0.1% DEPC-treated H2O).
- Centrifuged 4,000g for 2mins at RT.
- Discarded supernatant and repeated wash steps.
Pellet was resuspended in 50uL of 0.1% DEPC-treated H2O and stored @ -80oC in Ronit’s temporary box.