Monday 02/01/16

9:05 am – 7:30 pm, 8:15 pm – 11:40 pm

Imaging

  • testing control L4E24 embryos. stains look great.
  • compare to L7 embryos — no staining.
  • ran series removing S2 with toe-hold 5 bp. Looks excellent.

Troubleshooting

  • Gel tests
    • try to anneal probe and secondary on bench, no cells. Test if still bound on gel.
    • 5% PAGE gel. ran too long and gel was a bit too old
    • smears suggest partially degraded RNA slowing down template to various degrees
  • found the likely problem — 8 mM NaOH instead of 1 M NaOH in the RNA degradation step.
  • re-ran digestion, re-ran probe clean-up
  • re-ran hybridization

Ordering

  • S1rc + L7 common = GCGATGGTAGACGGCGTATGAATTCGGCAGAC GACCCGTCAGGATAGGCCCT

Microscope building prep

  • requesting quotes from MPB, Nikon, Coherent, and Newport

To do

  • write microscope justification
  • book flights back from San Fran
  • diagnostic gel
  • detailed labeling of samples
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