User:Madeline Hartman/Notebook/Dr. Hartings Lab/2012/11/27: Difference between revisions

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# 200 μL of cells were plated and spread with a sterile spreader
# 200 μL of cells were plated and spread with a sterile spreader
# The cells were incubated overnight at 37°C  
# The cells were incubated overnight at 37°C  
Cell transformation was unsuccessful


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Revision as of 13:02, 4 December 2012

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Entry title

Cell Transformation was performed
  1. BL21(DE3) cells thawed gently on ice
  2. The competent cells were mixed. 100 μL cells were added to three separate 14-ml BD Falcon polypropylene round bottom tubes.
  3. The three tubes were incubated on ice for 10 minutes, swirling gently ever 2 minutes
  4. 5 μL, 10 μL, and 15 μL of ligated DNA were added to one of the three tubes, and swirled gently
  5. The three tubes were incubated on ice for 30 minutes
  6. Each of the three tubes was heat-pulsed for 45 seconds in a 42°C water bath
  7. The three tubes were incubated on ice for 2 minutes
  8. 0.9 ml of LB medium was added to each of the three tubes and the tubes were incubated at 37 °C for 1 hour while shaking at 225 rpm.
  9. 200 μL of cells were plated and spread with a sterile spreader
  10. The cells were incubated overnight at 37°C

Cell transformation was unsuccessful


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