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Team:SDU-Denmark/labwork/Protocols/PCR
From 2012.igem.org
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<td><a href="https://2012.igem.org/Team:SDU-Denmark/labwork/Protocols/mutagen">Mutagenisis</a></td> | <td><a href="https://2012.igem.org/Team:SDU-Denmark/labwork/Protocols/mutagen">Mutagenisis</a></td> | ||
<td><a href="https://2012.igem.org/Team:SDU-Denmark/labwork/Protocols/pcrgelclean">PCR-,gel clean-up</a></td> | <td><a href="https://2012.igem.org/Team:SDU-Denmark/labwork/Protocols/pcrgelclean">PCR-,gel clean-up</a></td> | ||
| - | + | ||
</tr> | </tr> | ||
</table> | </table> | ||
Revision as of 17:03, 26 September 2012
| mRNA Isolation | PCR | Miniprep | Check Digest |
| 3A-Assembly | Colony-PCR | Transformation | Gel-electrophoresis |
| Reverse Transcriptase | Mutagenisis | PCR-,gel clean-up |
Polymerase Chain Reaction
Proof-reading PCR using Phusion Hot Start II
PCR program:
95°C for 30 seconds25-35 cycles: 98°C for 30 seconds Annealing temperature (5°C below primer melting temperature) 72°C for 15-30s/kb72°C for 5-10 minutes 4°C on hold
Non-proof-reading PCR using Taq Polymerase
5μl Dream Taq Buffer 5μl dNTP 1μl VF2 1μl VR Udtag 10μl fra primer stock og fortynd i 90μl oprenset vand 1,5μl Dream Taq Polymerase 1μl Template H2O up to 50μl PCR program:95°C for 2 minutes29 cycles: 95°C for 1 minut 55°C for 30 sek 72°C for 1min (1kb/min)72°C for 5 minutes 4°C on hold
