Blue:RNA-Seq Experiments:12022013

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Revision as of 19:57, 3 December 2013 by >B1lake (Created page with "== Human Nuclei C1 Experiment: 12-02-2013 == :'''FACS Sort''' :*DAPI stained human nuclei from Yun: :**Added ~1ul DAPI just prior to FACS sort on the Aria II (SCRM) :**Nucle...")
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Human Nuclei C1 Experiment: 12-02-2013

FACS Sort
  • DAPI stained human nuclei from Yun:
    • Added ~1ul DAPI just prior to FACS sort on the Aria II (SCRM)
    • Nuclei were maintained on ice
    • Added ~50ul of sort buffer containing 0.1% BSA to low-bind 1.5ml eppendorf tube for collection
    • Gated nuclei on DAPI positive and shifted gate slightly along the FSC axis to exclude the smallest nuclei
    • Sorted 150,000 nuclei into the collection tube (final volume of ~1200-1500ul)
    • Maintained on ice. Centrifuged 2000rpm at 4C for 10 minutes. Removed supernatant down to ~50ul. Resuspended pellet (not visible) using p1000 by pipetting about 5 times.
    • Counted nuclei using BioRad automatic cell counter: 1480 nuclei/ul