User:Karmella Haynes/Notebook/PcTF Genomics/2013/06/19

From OpenWetWare

< User:Karmella Haynes | Notebook | PcTF Genomics | 2013 | 06(Difference between revisions)
Jump to: navigation, search
(06/19/13)
Current revision (22:13, 26 June 2013) (view source)
(06/19/13)
 
(3 intermediate revisions not shown.)
Line 11: Line 11:
<!-- Precede finished items with a checkmark &#x2713; -->
<!-- Precede finished items with a checkmark &#x2713; -->
* Luc assay, time point 2
* Luc assay, time point 2
 +
* PcTF transfection for U2OS, outgrowth #2
 +
----
----
Line 54: Line 56:
|}
|}
 +
 +
 +
----
 +
'''PcTF transfection for U2OS, outgrowth #2'''
 +
 +
* Expanded transfected and mock-transfected cells from old 10 cm plates (6/15/13)
 +
* Used only samples 1 (+PcTF) and 4 (mock)
 +
* 1:10 passage into three new 10 cm plates labeled "1-A, 1-B, 1-C" (+PcTF) and "4-A, 4-B, 4-C" (mock)

Current revision

Pc-TF Genomics Main project page
Previous entry      Next entry


06/19/13

  • Luc assay, time point 2
  • PcTF transfection for U2OS, outgrowth #2



Luciferase activity assay - time point 2


Cell prep

  • Harvested cells
    • Seeded new dox+ plate with 1/2 of cells
    • Pelleted other half, resuspended in 2 mL FACS buffer


Assay reagents

  • Used the Biotium Steady-Luc Firefly HTS Assay Kit (same as 6/18/13)


Luc assay

  • Filtered 700 μL cells through strainer caps
  • Other steps same as 6/18/13


Cell counts

  • Wang lab's Accuri flow cytometer.
  • Set machine to read 20 uL of cells
  • Be sure to "clean" with water-run in between samples


Sample ID Gated count/ 20 μL   Cells/ 100 μL
sample 1 10,880 x5 = 54,400
sample 2 10,430 x5 = 52,150
sample 3 11,897 x5 = 59,485
sample 4 10,316 x5 = 51,580
sample 5 9,784 x5 = 48,920
sample 6 11,549 x5 = 57,745



PcTF transfection for U2OS, outgrowth #2

  • Expanded transfected and mock-transfected cells from old 10 cm plates (6/15/13)
  • Used only samples 1 (+PcTF) and 4 (mock)
  • 1:10 passage into three new 10 cm plates labeled "1-A, 1-B, 1-C" (+PcTF) and "4-A, 4-B, 4-C" (mock)



Personal tools