Difference between revisions of "Team:USMA-West Point/Experiments"

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<li>Warm up 50 mL DMEM (about 20 minutes in a 37 &#8451 water bath)</li>
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<li>Warm up 50 mL DMEM (about 20 minutes in a 37&deg; water bath)</li>
 
<li>Add 5 mL FSB (Fetal Bovine Serum)</li>
 
<li>Add 5 mL FSB (Fetal Bovine Serum)</li>
 
<li>Add 500 &#956;L Penstrep (Penicillin Streptomycin)</li>
 
<li>Add 500 &#956;L Penstrep (Penicillin Streptomycin)</li>

Revision as of 18:18, 13 September 2017

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USMA-West_Point

Experiments

Describe the research, experiments, and protocols you used in your iGEM project. These should be detailed enough for another team to repeat your experiments.

Please remember to put all characterization and measurement data for your parts on the corresponding Registry part pages.

What should this page contain?
  • Protocols
  • Experiments
  • Documentation of the development of your project
Inspiration


General Lab Etiquette
  • Lab coat, gloves, and goggles should be worn at all times
  • Spray everything that enters the hood with ethanol
  • Work past the windwall
  • Keep all lids face down
  • Bleach everything that was in contact with cells

HT22 Media Preparation
  1. Warm up 50 mL DMEM (about 20 minutes in a 37° water bath)
  2. Add 5 mL FSB (Fetal Bovine Serum)
  3. Add 500 μL Penstrep (Penicillin Streptomycin)
  4. Final concentrations should be 10% FBS and 1% Penstrep

Plating Cells
  1. Remove cells from liquid nitrogen
  2. Warm up cells in a water bath until a small crystal remains
  3. Pipette out media (1 mL) and place in a 10 mL Falcon tube
  4. Add 5 mL of media to the tube
  5. Centrifuge at 150 rcf for 5 min
  6. Vacuum out media without disturbing pellet
  7. Break pellet with 12 mL of media
  8. Distribute 2 mL into each well of a 6 well plate
  9. Incubate cells at 37 ℃ and 5% CO2

Changing Media
  1. Vacuum off media without touching the bottom of the plate
  2. Pipette 2 mL of media into each well
  3. Incubate cells at 37 ℃ and 5% CO2

Splitting Cells (Confirm with Rayonna)
  1. Vacuum out media
  2. Wash with PBS (Phosphate-buffered solution) and vacuum out
  3. Add trypsin to each well and incubate cells at 37 ℃ and 5% CO2 for 5 min
  4. Check for cell detachment, then remove trypsin/cell mixture

Freezing Cells