Difference between revisions of "Team:UCL/Experiments"

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<ol>
 
<li>(If using part from the distribution: resuspend the DNA in 10 ul of MiliQ water, making sure that it turns red. Wait 10 minutes before adding the DNA to cells)</li>
 
<li>(If using part from the distribution: resuspend the DNA in 10 ul of MiliQ water, making sure that it turns red. Wait 10 minutes before adding the DNA to cells)</li>
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2. Put a tube of NEB DH 5 alpha <i>E. coli</i> cells on ice and wait until they thaw completely. Divide the cells into 50 ul aliquotes. <br/>
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<li>Put a tube of NEB DH 5 alpha <i>E. coli</i> cells on ice and wait until they thaw completely. Divide the cells into 50 ul aliquotes. </li>
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3. Add 1 ul of plasmid DNA to 50 ul of cells. <br/>
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<li>Add 1 ul of plasmid DNA to 50 ul of cells. </li>
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4. Mix by carefully flicking the tube. Do not vortex or pipette in and out! <br/>
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<li>Mix by carefully flicking the tube. Do not vortex or pipette in and out! </li>
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5. Place the mixture on ice for 30 minutes. <br/>
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<li>Place the mixture on ice for 30 minutes. </li>
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6. Heat shock the cells at 42 °C for 30 seconds and immediately put on back on ice.<br/>
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<li>Heat shock the cells at 42 °C for 30 seconds and immediately put on back on ice.</li>
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7. Keep cells on ice for next 5 minutes. Do not mix. <br/>
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<li>Keep cells on ice for next 5 minutes. Do not mix. </li>
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8. Pipette 950 ul of SOC (or LB) media kept at room temperature into the mixture. <br/>
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<li>Pipette 950 ul of SOC (or LB) media kept at room temperature into the mixture. <br/>
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9. Incubate the mixture at 37 °C for 60 minutes <br/>
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<li>Incubate the mixture at 37 °C for 60 minutes </li>
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10. Plate 50 ul of cells to the plate with appropriate antibiotic and incubate overnight at 37 °C
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<li>Plate 50 ul of cells to the plate with appropriate antibiotic and incubate overnight at 37 °C</li>
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<br/><br/>
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Revision as of 22:13, 19 July 2015

Experiments & Protocols

Transformation protocol

  1. (If using part from the distribution: resuspend the DNA in 10 ul of MiliQ water, making sure that it turns red. Wait 10 minutes before adding the DNA to cells)
  2. Put a tube of NEB DH 5 alpha E. coli cells on ice and wait until they thaw completely. Divide the cells into 50 ul aliquotes.
  3. Add 1 ul of plasmid DNA to 50 ul of cells.
  4. Mix by carefully flicking the tube. Do not vortex or pipette in and out!
  5. Place the mixture on ice for 30 minutes.
  6. Heat shock the cells at 42 °C for 30 seconds and immediately put on back on ice.
  7. Keep cells on ice for next 5 minutes. Do not mix.
  8. Pipette 950 ul of SOC (or LB) media kept at room temperature into the mixture.
  9. Incubate the mixture at 37 °C for 60 minutes
  10. Plate 50 ul of cells to the plate with appropriate antibiotic and incubate overnight at 37 °C