10:30 A — 9:30 P
- mount non-post fixed bcd-STORM 647 labeled embryos
- post-fix remaining embryos
- No apparent bcd signal viewed on epi-fluorescence from STORM 647 label.
- Trying direct label with Alexa-647 anti-rabbit and Alexa-555-anti sheep (to cross react with hb anti-goat). Incubating 2.5 hours.
- Revise NSF proposal with recommendations from MB and send back for feedback.
- Primary antibodies don’t seem to react — no signal detected.
- Repeating antibody labeling using primaries at 1:100 at 4C overnight. Also test new laminin antibody. Using histone-GFP embryos as a labeling / detection control.
- Combinations Primaries:
C a-GFP + Rb a-Bcd + Gt a-Hb
C a-GFP + Rab a-Lamin + Gt a-Hb - Secondaries:
705 Dk a-C + A647 Dk a-Rb + A555 Dk a-Shp
705 Dk a-C + S647 Dk a-Rb + A563 Dk a-Gt
705 Dk a-C + A647 Dk a-Rb + A563 Dk a-Shp
705 Dk a-C + S647 Dk a-Rb + A555 Dk a-Gt - Bcd A-647 does work fine. Bcd 405-647 STORM dye also stains. Need lights off and 60x objective to see on Turnkey. STORM labeled Bcd 10x or more dimmer.
- Scan RSS literature feeds. Highlights: Cohesin binds PcG target genes whose expression goes up on Cohesin knockdown. Selectively targets paused genes (Gilmour +). Does cohesin increase long range enhancer repression by helping looping or help with chromatin compaction? Evidence for repressive loop formation?