User:Karmella Haynes/Notebook/PcTF Genomics/2014/03/17

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03/17/14

  • Mammalian cell culturing: Thawed KAH126 & KAH154 lines



Cell culture: Attempt to thaw PcTF stable lines
Request from Karsten Rippe, Heidelberg...

  1. U2OS Flp-in T-REx (parental cell line with no integration)
  2. U2OS Flp-in T-REx with hPCD-TF
  3. U2OS Flp-in T-REx with hPCD-RFP

Notes:

  • hPCD-TF = KAH126
  • hPCD-RFP = KAH154
  • First attempt at thawing KAH126 failed. Waited about 5 or so days to see growth.
  • Try again, but wait longer, see if small sub-population is viable & forms colonies

Procedure

  • Quick-thaw cells
  • Add 1 ml thawed cells to 5 ml 100% FBS
  • Pellet
  • Resuspend and grow in 4 mL 20% FBS, 1% P/S McCoy's 5A in T25 flasks
  • Grow at 37°C. Check every 5 days.



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