Team:Macquarie Australia/WetLab/Protocols/PCR

From 2014.igem.org

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<h3>PCR</h3>
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<h3>PCR</h3><br/>
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<h4>PCR Mixture: General recipe for PCR is as follows:</h4>
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<ul>
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<li>4 µL of 5x phusion buffer.</li>
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<li>0.4 µL dNTPs.</li>
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<li>0.6 µL of DMSO.</li>
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<li>0.2 µL of polymerase.</li>
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<li>11.8 µL of water.</li>
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</ul>
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<h4>To this mixture, add:</h4>
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<ul>
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<li>1 µL of forward primer.</li>
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<li>1 µL of reverse primer.</li>
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<li>1 µL of template.</li>
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</ul>
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<p>Total volume = 20 µL</p>
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<h4>PCR settings: A general program for PCR is as follows:</h4>
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<ul>
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<li>Initial denaturation at 98<sub>o</sub>C for 30seconds.</li>
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</ul>
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<p>Followed by 30 repeats of:</p>
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<ul>
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<li>Denaturation – 98oC for 10seconds.</li>
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<li>Annealing – 60<sub>o</sub>C for 10seconds.</li>
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<li>Extension – 72<sub>o</sub>C for 2minutes.</li>
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<li>Final extension – 72<sub>o</sub>C for 10minutes.</li>
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</ul>
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Revision as of 05:18, 16 October 2014

Protocols


PCR


PCR Mixture: General recipe for PCR is as follows:

  • 4 µL of 5x phusion buffer.
  • 0.4 µL dNTPs.
  • 0.6 µL of DMSO.
  • 0.2 µL of polymerase.
  • 11.8 µL of water.

To this mixture, add:

  • 1 µL of forward primer.
  • 1 µL of reverse primer.
  • 1 µL of template.

Total volume = 20 µL

PCR settings: A general program for PCR is as follows:

  • Initial denaturation at 98oC for 30seconds.

Followed by 30 repeats of:

  • Denaturation – 98oC for 10seconds.
  • Annealing – 60oC for 10seconds.
  • Extension – 72oC for 2minutes.
  • Final extension – 72oC for 10minutes.

Retrieved from "http://2014.igem.org/Team:Macquarie_Australia/WetLab/Protocols/PCR"