9:15 A — 6:45 P, 8:30P – 12:00A
- collect virgins for sim Espl[D] recombineering
- new cross, sna males x Sp/CyoZ; TM2/TM6 virgins
- clear MTDs and TM2/TM6s for virgin collection (peak emergence for latest bottles?)
- pulverize and refreeze Espl[D] and sim[D] flies for phenol extraction.
Confocal
- Attempting to flatten sim[D] samples with weight squeezed out mounting media = one big air bubble. sample mostly ruined and very few sim[D] embryos left to work with.
- imaged anyway. Possibly slightly wider, maybe less sharp, maybe a temporal thing before snail appears.
- some sim[D] lines have non-uniform gastrulation (with and without sim expression, though maybe more common without).
- Image Psc embryo for staining control for SCM. copy SCM images. Still need to write up summary.
Shouldn’t have frozen SCM flies, could have happily continued in 18C for another week. Now it will be very difficult to screen specifically for early stages to follow up mutant work. Can always remake this line but a stupid waste of time. Oh well.
Probe making
- glycogen arrived. Percipitate and freeze samples.
- Vortex mix after ethanol added is very important
- 2 mL is a little small for the amount of EtOH added with 600 uL aliquots. Should recompute for 500 or 400 uL of probe per 200 mL tube… This protocol also uses A LOT of glycogen. Will probably need to order more…
- 2 tubes not sufficiently mixed partially frozen. Thawing and mix and refreezing O/N at 20C (preferred method according to protocol).
analyze inv data
- issues with background from 647 channel. Can filter out but shouldn’t be an issue if the lasers would work right. (see post).
- Good 647 coverage in sections imaged so far. very suggestive colocalization structures one silenced (tou) marked loci examined so far.
Finish first BJ draft.
- First read end to end with introduction finished. Still need to write conclusion / perspective.
- Current draft is 8000 words main text. 5 pages is closer to 4000 (probably less with substantial figures).
- skipping perspective/conclusion for now. Better to get feedback from ML get focus defined, and then go back and write, meanwhile the intro will stand.
- Sent to ML with figures.