From 2009.igem.org
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Protocols
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Ligation
(estimated time: 20 min + 12-16 hours overnight incubation)
- (For every ligation)
- Add 50 ng of vector
- Add
- Heat DNA mix at 65°C for 5 min for DNA denaturation
- Add 1 µl of T4 Ligase buffer
- Add 1 µl of T4 Ligase
- 10 µl final volume
- Incubate at 16°C overnight
- Then, ligation can be conserved at 4°C or can be transformed
- Before transformation you have to inactivate T4 Ligase:
- Heat ligation at 65°C for 10 min.
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- Roche T4 Ligase
- 10X Roche T4 Ligase Buffer
- ddH2O
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