8:30a – 6:00p, 8:00p – 11:10p
T7 plate reaction planning (?)
- NEB Quick High Yield T7
- template = 10 uL of T7 template (~30-120 ng/uL from spot check)
- 10 uL NTP buffer mix + 1uL (1/2x) T7 polII mix + 10 uL template (~1 ug)
- 820 uL NTP buffer mix + 82 uL T7 enzyme mix.
- supplemental NTP buffer mix (400 ul) = 40 uL 25mM rNTPs + 40 uL RNasin + 80 uL 10x buffer + 240 uL ddH2O.
- incubating at 37C, from 2:30P.
STORM analysis
- insightM analysis giving weird results — loaded molecule lists back into STORMfinder don’t hit most of the molecules. The frame by frame finder does much better. I’ve seen this before and I can’t remember what causes the error.
Review writing
Review feedback
- Why use Markov Models
- How we are going to use Markov models: cartoon, equation, evaluate with computer algebra, interpret analytical solution
- will be illustrated with examples from literature, examples should be clear enough to build on
- Need to talk through how to build the Matrix
- Don’t need extra algebra in the result
- Mention briefly model approaches in intro.
Additional Goals
- write paragraph on Thermodynamic gene reg models in development
- write conclusion paragraph
New STORM buffer
- made new glox
- making aliquots of Cyclooctatetraene
- molecular weight 104.15. 200 mM stock. 104 g in 1 L is 1M. 200 mg in 1 mL is 2M. 20 mg in 1 mL DMSO is 200 mM.
- made 14 aliquots from 1 mL of 200 mM COT stock
- sample 2 dots look good.
- hal not talking to 405 laser, unable to do STORM
Things for discussion with XZ on Thursday
- keeping feducials attached to sample. (check cross-linked beads tonight/tomorrow)
Banff
- write to participants, final reminder / planning.