Matt:LabNotes/2015-6-5
Jump to navigation
Jump to search
RNA FISH: Probe Resuspension and Dye Coupling
Probe Resuspension
- The 5' amino modified (separated by C6) 48-probe set ordered by Dan:
- GAD1
- Already resuspended
- FOXP2
- Already resuspended
- CUX2
- Already resuspended
- SLC6A1
- ADARB2
- SATB2
- KIT
- GAD1
- Pick 2-3 genes and resuspend if needed
- Dr. Zhang suggested SLC6A1 and SATB2
- Table of ordered plates
- SLC6A1: Plate=CUX2-SLC6A1-20150128 Wells=E1-H12
- SATB2: Plate=ADARB2-SATB2-20150128 Wells=E1-H12
- Gene information with CA12kNov14_V4 efficiencies
- Table of ordered plates
- Resuspend each oligo in 15 uL nuclase-free H2O (1 mM)
- Take 2 uL from each oligo well and combine them in a single 1.5 mL tube (total concentration 1 mM)
- Store the remaining oligos at -20C
- Add 432 uL nuclase-free, diluting 10:1, for final concentration of 100 uM
- Technically closer to 5:1 dilution but final result is closer to 10:1 because starting concentration is actually lower...
- Check concentration in nanodrop at 1:1, 5:1, and 10:1 dilutions for accuracy
Dye Coupling
Dan's best practice dye coupling protocol
Generic dye coupling protocol