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Cloak
11 complete 6/23: Seed cells 12 complete 6/24: Transfect hEF1a:eYFP into HEK cells 13 complete 6/26 (4.30pm): Flow Cytometer
  • Determine optimal Lipofectamine LTX volume (24 well)
  • Determine transfection efficiency (quantitatively)

...

Cloak
22 incomplete 7/2: Seed cells 23 incomplete 7/3: Transfect hEF1a:eYFP and hEF1a:mKate into HEK cells 24 incomplete 7/5 (__pm): Flow Cytometer

PLATE:

 

/////////
Y1
  • _uL complete media
  • _uL hEF1a:eYFP
  • _uL Lipofectamine LTX
(Y+R)1 Y2
  • _uL complete media
  • _uL hEF1a:eYFPmKate
  • _uL Lipofectamine LTX
//////   

-ve control

  • dummy DNA
 
Y2
  • _uL complete media
  • _uL hEF1a:eYFP
  • _uL Lipofectamine LTX
(Y+R)Y42
  • _uL complete media
  • _uL hEF1a:eYFPmKate
  • _uL Lipofectamine LTX
//////   

-ve control

  • dummy DNA
R1
  • _uL complete media
  • _uL hEF1a:mKate
  • _uL Lipofectamine LTX
(Y+R)3 R2
  • _uL complete media
  • _uL hEF1a:mKate
  • _uL Lipofectamine LTX
//////   

-ve/-ve control

  • cells (no action)
 
R2
  • _uL complete media
  • _uL hEF1a:mKate
  • _uL Lipofectamine LTX
(Y+R)R4
  • _uL complete media
  • _uL hEF1a:mKate
  • _uL Lipofectamine LTX
 /////////  

-ve/-ve control

  • cells (no action)

 

 

Toggle Cloak
PRACTICE #3: Fluorescent protein under inducible promoter

Cloak
14 incomplete 7/_: Seed cells 15 incomplete 7/_: Transfect TRE:mKate, hEF1a:rtTA and hEF1a:eYFP (transfection marker) into HEK cells 16 incomplete 7/_: Add DOX 18 incomplete 7/_ (__pm): Flow Cytometer

 

Y1
  • _uL complete media
  • _uL hEF1a:eYFP
  • _uL Lipofectamine LTX
(Y+R)1
  • _uL complete media
  • _uL hEF1a:mKate
  • _uL Lipofectamine LTX
   

-ve control

  • dummy DNA
Y2
  • _uL complete media
  • _uL hEF1a:eYFP
  • _uL Lipofectamine LTX
(Y+R)2
  • _uL complete media
  • _uL hEF1a:mKate
  • _uL Lipofectamine LTX
   

-ve control

  • dummy DNA
R1
  • _uL complete media
  • _uL hEF1a:mKate
  • _uL Lipofectamine LTX
(Y+R)3
  • _uL complete media
  • _uL hEF1a:mKate
  • _uL Lipofectamine LTX
   

-ve/-ve control

  • cells (no action)
R2
  • _uL complete media
  • _uL hEF1a:mKate
  • _uL Lipofectamine LTX
(Y+R)4
  • _uL complete media
  • _uL hEF1a:mKate
  • _uL Lipofectamine LTX
   

-ve/-ve control

  • cells (no action)

Toggle Cloak
PRACTICE #4**: Co-transfection (2 fluorescent proteins under constitutive & inducible promoters)

Cloak
29 incomplete 7/_: Seed cells 30 incomplete 7/_: Transfect hEF1a:eYFP, TRE:mKate and hEF1a:rtTA** into HEK cells 31 incomplete 7/_: Add DOX 32 incomplete 7/_ (__pm): Flow Cytometer

 

Y1
  • _uL complete media
  • _uL hEF1a:eYFP
  • _uL Lipofectamine LTX
(Y+R)1
  • _uL complete media
  • _uL hEF1a:mKate
  • _uL Lipofectamine LTX
   

-ve control

  • dummy DNA
Y2
  • _uL complete media
  • _uL hEF1a:eYFP
  • _uL Lipofectamine LTX
(Y+R)2
  • _uL complete media
  • _uL hEF1a:mKate
  • _uL Lipofectamine LTX
   

-ve control

  • dummy DNA
R1
  • _uL complete media
  • _uL hEF1a:mKate
  • _uL Lipofectamine LTX
(Y+R)3
  • _uL complete media
  • _uL hEF1a:mKate
  • _uL Lipofectamine LTX
   

-ve/-ve control

  • cells (no action)
R2
  • _uL complete media
  • _uL hEF1a:mKate
  • _uL Lipofectamine LTX
(Y+R)4
  • _uL complete media
  • _uL hEF1a:mKate
  • _uL Lipofectamine LTX
   

-ve/-ve control

  • cells (no action)