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Team:SDU-Denmark/labwork/Protocols/colpcr
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<td><a href="https://2012.igem.org/Team:SDU-Denmark/labwork/Protocols/revtrans">Reverse Transcriptase</a></td> | <td><a href="https://2012.igem.org/Team:SDU-Denmark/labwork/Protocols/revtrans">Reverse Transcriptase</a></td> | ||
| - | <td><a href="https://2012.igem.org/Team:SDU-Denmark/labwork/Protocols/mutagen"> | + | <td><a href="https://2012.igem.org/Team:SDU-Denmark/labwork/Protocols/mutagen">Mutagenesis</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> | ||
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Latest revision as of 21:09, 26 September 2012
| mRNA Isolation | PCR | Miniprep | Check Digest |
| 3A-Assembly | Colony-PCR | Transformation | Gel-electrophoresis |
| Reverse Transcriptase | Mutagenesis | PCR-,gel clean-up |
Colony PCR
Transfer few cells from a colony to the bottom of a PCR tube. Add 100uL of sterilized water to the tube and microwave it at max effect for 2 mins to destroy the cells.Use 2uL of the colony DNA template suspension in the following PCR mix. PCR Mix:
5μl Dream Taq Buffer 1μl dNTP 1μl Primer1 (VF2) 1μl Primer2 (VR) 3μl 25mM MgCl2 1μl Dream Taq Polymerase 2μl Template suspension 36μl sterile destilled H2O
PCR Program:
95°C for 2minutes30 cycles: 95°C for 1 minut melting temp -5°C for 30seconds 72°C for 1 minute/kb72°C for 10 minute 4°C on hold
