User:Karmella Haynes/Notebook/PcTF Genomics/2013/06/15: Difference between revisions

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(Autocreate 2013/06/15 Entry for User:Karmella_Haynes/Notebook/PcTF_Genomics)
 
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==mm/dd/yy==
==06/15/13==
<!-- Precede finished items with a checkmark &#x2713; -->
<!-- Precede finished items with a checkmark &#x2713; -->
* Line item 1
* Gal4-EED/luc Luciferase ICC - fixation, permeabilization, primary ab staining
* Gal4-EED/ luc dox induction - shipment 2 cells
* PcTF transfection for U2OS, outgrowth #1
 


----
----
'''Line item 1'''<br>
'''Gal4-EED/luc Luciferase ICC'''<br>
> Samples
 
Samples (Shipment 1 cells)
# 0 μg/mL dox
# 0.1
# 0.2
# 0.5
# 1.0
# 2.0 (induced on 6/02/13, 13 days)
 
 
* Followed standard protocol
* Harvested cells from one 6-well plate (split plate #2 for further analyses)
* At the primary antibody staining step, keep samples and reagents on ice overnight in the cold room


{| class="wikitable" border="0" cellspacing="3" <!-- Rxn. table -->
|-valign="top"
| <u>Reagent</u> || <u>Volume</u>
|-
| reagent 1 || # μL
|-
| reagent 2 || #
|-
| reagent 3 || #
|-
| reagent 4 || #
|-
| dH<sub>2</sub>O || #
|-
| &nbsp; || # μL
|}


--> Reaction conditions
----
'''Gal4-EED/luc dox induction'''<br>
 
Samples (Shipment 2 cells); new 6-well plate...
# 0 μg/mL dox
# 0.1
# 0.2
# 0.5
# 1.0
# 2.0 (induced on 6/02/13, 13 days)
 
* For this set, test earlier time points and try luc activity assay
 
 
 
----
'''PcTF transfection for U2OS, outgrowth #1'''
* 6-well plates: 1-3 = +PcTF, 4-6 = mock (lipo only)
* Expand cells from 6-well plates in individual 10 cm plates (10 mL medium). 6 10 cm plates total.
* Use standard growth medium.
* Use all cells. Do not dilute.




Revision as of 20:11, 26 June 2013

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06/15/13

  • Gal4-EED/luc Luciferase ICC - fixation, permeabilization, primary ab staining
  • Gal4-EED/ luc dox induction - shipment 2 cells
  • PcTF transfection for U2OS, outgrowth #1


Gal4-EED/luc Luciferase ICC

Samples (Shipment 1 cells)

  1. 0 μg/mL dox
  2. 0.1
  3. 0.2
  4. 0.5
  5. 1.0
  6. 2.0 (induced on 6/02/13, 13 days)


  • Followed standard protocol
  • Harvested cells from one 6-well plate (split plate #2 for further analyses)
  • At the primary antibody staining step, keep samples and reagents on ice overnight in the cold room


Gal4-EED/luc dox induction

Samples (Shipment 2 cells); new 6-well plate...

  1. 0 μg/mL dox
  2. 0.1
  3. 0.2
  4. 0.5
  5. 1.0
  6. 2.0 (induced on 6/02/13, 13 days)
  • For this set, test earlier time points and try luc activity assay


PcTF transfection for U2OS, outgrowth #1

  • 6-well plates: 1-3 = +PcTF, 4-6 = mock (lipo only)
  • Expand cells from 6-well plates in individual 10 cm plates (10 mL medium). 6 10 cm plates total.
  • Use standard growth medium.
  • Use all cells. Do not dilute.



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