User:Karmella Haynes/Notebook/PcTF Genomics/2013/06/10: Difference between revisions

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(Autocreate 2013/06/10 Entry for User:Karmella_Haynes/Notebook/PcTF_Genomics)
 
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==mm/dd/yy==
==06/10/13==
<!-- Precede finished items with a checkmark &#x2713; -->
<!-- Precede finished items with a checkmark &#x2713; -->
* Line item 1
* Split cells: Gal4-EED/luc shipment 1 and shipment 2 T-75 flasks (1:5)
* Split dox induced cells
 


----
----
'''Line item 1'''<br>
'''Gal4EED/luc test'''<br>
> Samples
> Dox dose response test<br>
 
 
6-well plates for luc activity assays & flow cytometry, etc.
* 6-well plate (originally plated 5/29/13), shipment 1 cells
* Split 1:4 into 2 new 6-well plates: resuspend detached cells in a final volume of 4 mL puro+ medium, add 1 mL cells to 3mL fresh medium +proper amount of dox


{| class="wikitable" border="0" cellspacing="3" <!-- Rxn. table -->
{| class="wikitable" border="0" cellspacing="5" <!-- Cell plating table -->
|-valign="top"
| <u>Reagent</u> || <u>Volume</u>  
|-
|-
| reagent 1 || # μL
| <u>Wells</u> || <u>dox μg/mL</u> || <u>Volume dox (1 mg/mL) in 4.0 mL medium</u>
|-
|-
| reagent 2 || #
| 1            || 0        || ---     
|-
|-
| reagent 3 || #
| 2            || 0.1        || 0.5 μL
|-
|-
| reagent 4 || #
| 3            || 0.2        || 1.0 μL
|-
|-
| dH<sub>2</sub>O || #
| 4            || 0.5        || 2.0 μL
|-
|-
| &nbsp; || # μL
| 5            || 1.0        || 4.0 μL
|-
| 6            || 2.0        || 8.0 μL  
|}
|}


--> Reaction conditions
 
24-well plate for microscopy:
* Use leftover suspension from 6-well plating
* For each sample (1-6) add 0.5 mL suspension to 0.5 mL fresh medium +proper amount of dox in 4 wells
* Cells will be fixed and stained directly in the plate, following David Dreher's protocol.




Revision as of 20:19, 12 June 2013

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06/10/13

  • Split cells: Gal4-EED/luc shipment 1 and shipment 2 T-75 flasks (1:5)
  • Split dox induced cells


Gal4EED/luc test
> Dox dose response test


6-well plates for luc activity assays & flow cytometry, etc.

  • 6-well plate (originally plated 5/29/13), shipment 1 cells
  • Split 1:4 into 2 new 6-well plates: resuspend detached cells in a final volume of 4 mL puro+ medium, add 1 mL cells to 3mL fresh medium +proper amount of dox
Wells dox μg/mL Volume dox (1 mg/mL) in 4.0 mL medium
1 0 ---
2 0.1 0.5 μL
3 0.2 1.0 μL
4 0.5 2.0 μL
5 1.0 4.0 μL
6 2.0 8.0 μL


24-well plate for microscopy:

  • Use leftover suspension from 6-well plating
  • For each sample (1-6) add 0.5 mL suspension to 0.5 mL fresh medium +proper amount of dox in 4 wells
  • Cells will be fixed and stained directly in the plate, following David Dreher's protocol.



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