User:Karmella Haynes/Notebook/PcTF Genomics/2013/07/04: Difference between revisions
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|style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;">Pc-TF Genomics</span> | |style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;">Pc-TF Genomics</span> | ||
|style="background-color: #F2F2F2" align="center"| | |style="background-color: #F2F2F2" align="center"|[[File:Report.png|frameless|link={{#sub:{{FULLPAGENAME}}|0|-11}}]][[{{#sub:{{FULLPAGENAME}}|0|-11}}|Main project page]]<br />{{#if:{{#lnpreventry:{{FULLPAGENAME}}}}|[[File:Resultset_previous.png|frameless|link={{#lnpreventry:{{FULLPAGENAME}}}}]][[{{#lnpreventry:{{FULLPAGENAME}}}}{{!}}Previous entry]] }}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]][[File:Resultset_next.png|frameless|link={{#lnnextentry:{{FULLPAGENAME}}}}]]}} | ||
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==07/04/13== | ==07/04/13== | ||
<!-- Precede finished items with a checkmark ✓ --> | <!-- Precede finished items with a checkmark ✓ --> | ||
* | * Gal4-EED/luc time/dose - luc assay & cell counts | ||
* Happy July 4th! :) | * Happy July 4th! :) | ||
* Gal4-EED/luc Dox induction #2 | |||
---- | ---- | ||
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<u>Assay reagents</u> | <u>Assay reagents</u> | ||
* Used the Biotium Steady-Luc Firefly HTS Assay Kit (same as [http://openwetware.org/wiki/User:Karmella_Haynes/Notebook/PcTF_Genomics/2013/ | * Used the Biotium Steady-Luc Firefly HTS Assay Kit (same as [http://openwetware.org/wiki/User:Karmella_Haynes/Notebook/PcTF_Genomics/2013/06/18 6/18/13]) | ||
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* Samples loaded in triplicate (by columns) | * Samples loaded in triplicate (by columns) | ||
* Included luc buffer + FACS-buffer "blank" sample (well D1) | * Included luc buffer + FACS-buffer "blank" sample (well D1) | ||
* Other steps same as [http://openwetware.org/wiki/User:Karmella_Haynes/Notebook/PcTF_Genomics/2013/ | * Other steps same as [http://openwetware.org/wiki/User:Karmella_Haynes/Notebook/PcTF_Genomics/2013/06/18 6/18/13] | ||
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| sample 5 || 16,545 || x5 = || 82,725 | | sample 5 || 16,545 || x5 = || 82,725 | ||
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| sample 6 || || x5 = || | | sample 6 || 17,885 || x5 = || 89,425 | ||
|} | |} | ||
---- | |||
'''Gal4-EED/luc Dox induction #2'''<br> | |||
* Lower range worked very well for trail #1! Saw ranges in expression for 1.0 - 10<sup>-4</sup> μg/mL doc after 4 days | |||
* Repeat induction and test after 48 hours | |||
# 1.0 μg/mL dox | |||
# 10<sup>-1</sup> | |||
# 10<sup>-2</sup> | |||
# 10<sup>-3</sup> | |||
# 10<sup>-4</sup> | |||
# 0 | |||
Addition of serial dilutions of dox to cells | |||
* Start with 3.5 mL growth medium in cell culture wells. Remove some growth medium from cells where necessary. | |||
* Set up the following serial dilutions (using appropriate growth medium) in sterile 1 mL tubes... | |||
{| {{table}} | |||
|- | |||
| || tube 1 || 2 || 3 || 4 || 5 || 6 | |||
|- | |||
| 1 mg/mL dox (μL) || 8.0 || --- || --- || --- || --- || --- | |||
|- | |||
| mixture from prior tube (μL) || --- || 100 || 100 || 100 || 100 || --- | |||
|- | |||
| +growth med. (μL) || 1000 || 900 || 900 || 900 || 900 || --- | |||
|- | |||
| Final [dox] (μg/mL) in 4 mL med. || 1.0 || 10<sup>-1</sup> || 10<sup>-2</sup> || 10<sup>-3</sup> || 10<sup>-4</sup> || 0 | |||
|} | |||
Latest revision as of 22:58, 26 September 2017
Pc-TF Genomics | Main project page Previous entry Next entry | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
07/04/13
Luciferase activity assay - time point: 4 days
Gal4-EED/luc Dox induction #2
Addition of serial dilutions of dox to cells
|