User:Karmella Haynes/Notebook/PcTF Genomics/2014/11/15: 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"|<html><img src="/images/9/94/Report.png" border="0" /></html> [[{{#sub:{{FULLPAGENAME}}|0|-11}}|Main project page]]<br />{{#if:{{#lnpreventry:{{FULLPAGENAME}}}}|<html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>[[{{#lnpreventry:{{FULLPAGENAME}}}}{{!}}Previous entry]]<html>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;</html>}}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]]<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html>}}
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# KAH126/MV2
# KAH126/MV2
# KAH126/MV2
# mock
# no transfection
# no transfection


* Grow overnight, look for RFP next day
* Grow overnight, look for RFP next day
* Next day: harvest transfected cells and use 1/2 (5.0E5) for 2 different plates
* Next day: harvest transfected cells and use 1/2 (5.0E5) for 2 different plates
** 24-well plate - ~2.1E5 cells/well
** 24-well plate - ~2.1E4 cells/well
** 96-well plate -  
** 96-well plate - ~5.2E4 cells/well
 


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Latest revision as of 00:31, 27 September 2017

Pc-TF Genomics Main project page
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11/15/14

  • Transfection: KAH126/MV2 into U2OS


Transfection: KAH126/MV2 into U2OS
Strategy: instead of trying to increase transfection efficiency, do a "quasi-subcloning" where transfected cells are separated into smaller wells, and wells showing highest RFP will be selected, pooled, and re-plated.

  • 6-well plate seeded on 11/14/14 (~2.5E5 cells/well)
  • Change medium to p/s-free before transfection (4 mL per well)
  • Use standard Lipofectamine LTX procedure w/ 2000 ng plasmid
  1. KAH126/MV2
  2. mock
  3. no transfection
  • Grow overnight, look for RFP next day
  • Next day: harvest transfected cells and use 1/2 (5.0E5) for 2 different plates
    • 24-well plate - ~2.1E4 cells/well
    • 96-well plate - ~5.2E4 cells/well



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