Difference between revisions of "Team:Stony Brook/Notebook/Cancer-W14"

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<h4> Toggle 2 Gel Construct using Phire Polymerase </h4>
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<h4> PCR Settings Template: </h4>
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 +
<ul>
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<table>
 +
<tr>
 +
<th> Phase </th>
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<th> Temperature (°C) </th>
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<th> Time (sec)</th>
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</tr>
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<tr>
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<td> Initial Denaturation </td>
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<td> 98 </td>
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<td> 60s </td>
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</tr>
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<tr>
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<td> Denaturation </td>
 +
<td> 98 </td>
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<td> 20s </td>
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</tr>
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<tr>
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<td> Annealing </td>
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<td> Ranged from 72-60 degrees for each tube. Did a different annealing temperature for each PCR. </td>
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<td> 10s </td>
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</tr>
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<tr>
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<td> Extension </td>
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<td> 72 </td>
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<td> # </td>
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</tr>
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<tr>
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<td> Final Extension </td>
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<td> 72 </td>
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<td> # </td>
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</tr>
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</table>
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<h6> Ran 32 cycles </h6>
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</ul>
  
 
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Revision as of 02:48, 12 October 2016

Week 14: 9/26 - 10/2

Week 14: 9/26 - 10/2



9/26



9/27



9/28



9/29

Toggle 2 Gel Construct using Phire Polymerase

PCR Settings Template:

    Phase Temperature (°C) Time (sec)
    Initial Denaturation 98 60s
    Denaturation 98 20s
    Annealing Ranged from 72-60 degrees for each tube. Did a different annealing temperature for each PCR. 10s
    Extension 72 #
    Final Extension 72 #
    Ran 32 cycles


9/30



10/1



10/2

Transformations

  1. Thaw TSS cells on ice
  2. Add 2ul of prepped plasmid. Pipette to mix
  3. Sit for 30 minutes on ice
  4. Incubate cells for 30s at 43°C
  5. Incubate on ice for 2 minutes
  6. Add 1ml of 50C
  7. Incubate for 30 minutes at 37°C. Shake
  8. Spread 100-300ul on plates
  9. Grow overnight at 37°C
  10. Save the rest as liquid cultures

Plates Used:

  • Plate 1: pEPGAP
  • Plate 2: pEPGAP/CRISBU
  • Plate 3: pEPGAP-P
  • Plate 4:
  • Plate 5: pSBIA2/T1/T2
  • Plate 6: pSBIA2
  • Plate 7: pSBIA2-P
  • Plate 8: