Team:British Columbia/Notebook/Protocols/transformation

From 2014.igem.org

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<li> Add 1µL ligation mix to thawed cells and incubate for 30 minutes on ice.
<li> Add 1µL ligation mix to thawed cells and incubate for 30 minutes on ice.
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<li>Heat shock in water bath/heat block for 60 seconds and put back on ice for two minutes.
<li>Heat shock in water bath/heat block for 60 seconds and put back on ice for two minutes.
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<li> Add 400uL LB Broth and incubate at 37C for 2 hours.
<li> Add 400uL LB Broth and incubate at 37C for 2 hours.
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<li> Spread plate entire 500µL.</li>
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<li> Spread plate entire 500µL.
 
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<li>Turn off the flame </li>
 
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<li>Turn off the flame.</li>
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<li> Incubate overnight (<18hrs) at 37°C.</li>
<li> Incubate overnight (<18hrs) at 37°C.</li>

Revision as of 01:07, 18 October 2014

2014 UBC iGEM

Transformation


Supplies:

  • Competent cells in 100μL aliquots.
  • Water bath/heat block at 42°C.
  • Ice
  • LB Broth
  • LB agar plates (with appropriate selection)

Steps:

  1. Remove competent cells (100uL aliquots) from -80°C and thaw on ice.
    • Note, aliquots can be thawed GENTLY by hand if time is an issue*

  2. Add 1µL ligation mix to thawed cells and incubate for 30 minutes on ice.
  3. Heat shock in water bath/heat block for 60 seconds and put back on ice for two minutes.
    • Remember to turn off the water bath.
    • The following steps must be done near a flame.
    • Don’t forget a growth control, especially if you aren’t screening using antibiotics.

  4. Add 400uL LB Broth and incubate at 37C for 2 hours.
    • Note, 1 hour is also sufficient

  5. Spread plate entire 500µL.

  6. Turn off the flame.

  7. Incubate overnight (<18hrs) at 37°C.


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