User:Behzad Damadzadeh/Notebook/PcTF Subcloning in E-coli/2013/06/17

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Transfer [http://openwetware.org/wiki/User:Behzad_Damadzadeh/Notebook/PcTF_Subcloning_in_E-coli/2013/06/10 Transfected Cells] from 6 well plate to 10cm cell culture dishes.
Transfer [http://openwetware.org/wiki/User:Behzad_Damadzadeh/Notebook/PcTF_Subcloning_in_E-coli/2013/06/10 Transfected Cells] from 6 well plate to 10cm cell culture dishes.
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Target cell concentrations are: 1*10
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Target cell concentrations are: 1x10<suP>6</sup>, 1x10<suP>5</sup>, 1x10<suP>4</sup>, 1x10<suP>3</sup>
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*Add 2mL Trypsin to each of the wells 1 and 2 for 4 minutes.
*Add 2mL Trypsin to each of the wells 1 and 2 for 4 minutes.
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* Add 3.5mL media to neutralize Trypsin.
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*Add 3.5mL media to neutralize Trypsin.
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*Add 6mL media to the 4ml of well 1.
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*Transfer all 10mL of well 1 to 10cm plate to have 1x10<suP>6</sup>.
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*Add 16mL media to the 4mL of well 2.
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*Take 2mL of cells, dilute with 8mL of media and transfer to 10cm plate 1x10<suP>5</sup>.
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*Take 1mL of cells in well 2, dilute with 9mL of media and transfer to 10cm plate 1x10<suP>4</sup>.
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*Take 0.5mL of cells in well 2, dilute in 9.5mL of media and transfer to 10cm plate 1x10<suP>3</sup>.

Revision as of 18:57, 23 July 2013

PcTF Subcloning in E. coli Main project page
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06/17/2013

Prepare Transfected Cells for Hygromycine Treatment

Transfer Transfected Cells from 6 well plate to 10cm cell culture dishes. Target cell concentrations are: 1x106, 1x105, 1x104, 1x103


  • Add 2mL Trypsin to each of the wells 1 and 2 for 4 minutes.
  • Add 3.5mL media to neutralize Trypsin.
  • Add 6mL media to the 4ml of well 1.
  • Transfer all 10mL of well 1 to 10cm plate to have 1x106.
  • Add 16mL media to the 4mL of well 2.
  • Take 2mL of cells, dilute with 8mL of media and transfer to 10cm plate 1x105.
  • Take 1mL of cells in well 2, dilute with 9mL of media and transfer to 10cm plate 1x104.
  • Take 0.5mL of cells in well 2, dilute in 9.5mL of media and transfer to 10cm plate 1x103.
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