"
Team:Macquarie Australia/WetLab/Protocols/PCR
From 2014.igem.org
(Difference between revisions)
| (One intermediate revision not shown) | |||
| Line 22: | Line 22: | ||
<li><a href="https://2014.igem.org/Team:Macquarie_Australia/WetLab/Protocols/SDSPAGE">SDS-PAGE</a></li> | <li><a href="https://2014.igem.org/Team:Macquarie_Australia/WetLab/Protocols/SDSPAGE">SDS-PAGE</a></li> | ||
<li><a href="https://2014.igem.org/Team:Macquarie_Australia/WetLab/Protocols/Heat">Heat Shock</a></li> | <li><a href="https://2014.igem.org/Team:Macquarie_Australia/WetLab/Protocols/Heat">Heat Shock</a></li> | ||
| + | <li><a href="https://2014.igem.org/Team:Macquarie_Australia/WetLab/Protocols/Operons">Induction of Operons</a></li> | ||
</ul> | </ul> | ||
</div> | </div> | ||
| Line 47: | Line 48: | ||
<h4>PCR settings: A general program for PCR is as follows:</h4> | <h4>PCR settings: A general program for PCR is as follows:</h4> | ||
<ul> | <ul> | ||
| - | <li>Initial denaturation at 98< | + | <li>Initial denaturation at 98<sup>o</sup>C for 30seconds.</li> |
</ul> | </ul> | ||
<h4>Followed by 30 repeats of:</h4> | <h4>Followed by 30 repeats of:</h4> | ||
<ul> | <ul> | ||
| - | <li>Denaturation – | + | <li>Denaturation – 98<sup>o</sup>C for 10seconds.</li> |
| - | <li>Annealing – 60< | + | <li>Annealing – 60<sup>o</sup>C for 10seconds.</li> |
| - | <li>Extension – 72< | + | <li>Extension – 72<sup>o</sup>C for 2minutes.</li> |
| - | <li>Final extension – 72< | + | <li>Final extension – 72<sup>o</sup>C for 10minutes.</li> |
</ul> | </ul> | ||
Latest revision as of 13:10, 17 October 2014
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.
