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Team:ETHZ Basel/Biology/Protocols
From 2010.igem.org
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== ligation == | == ligation == | ||
| - | 10 ng storage plasmid | + | <br>10 ng storage plasmid |
| - | 10 times more insert | + | <br>10 times more insert |
| - | 1 ul ligase buffer (10x T4) | + | <br>1 ul ligase buffer (10x T4) |
| - | fill up water to 10 ul | + | <br>fill up water to 10 ul |
== transformation == | == transformation == | ||
Revision as of 13:37, 5 August 2010
Contents |
PCR
10 ul 5X Buffer
1 ul dNTP's
2 ul Primers (10 fold diluted)
1 ul Template
0.5 ul high fidelity polymerase
35.5 ul water
98°C 30s
98°C 30s
55°C 30s
72°C 60s
52°C 5min
4°C
35 cycles
ligation
10 ng storage plasmid
10 times more insert
1 ul ligase buffer (10x T4)
fill up water to 10 ul
transformation
chemical:
isolation of plasmids
according to the protocol of NucleoSpin Plasmid (Macherey-Nagel)
agarose gel
1% agarose gel 0.5 g agarose 50 ml 1X TAE heat up with microwave, let cool down a bit 1 ul EtBr pour
