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Team:Groningen/protocols/Ligation
From 2013.igem.org
Ligation Reaction
Materials:
- MQ water
- 1.5ml tubes
- 10x T4 Ligation buffer
- T4 Ligase
- Insert
- Vector
Reaction mixture:
| Component | 20µl | Final concentration |
|---|---|---|
| MilliQ water | up to 20µl | |
| 10 ligation buffer | 2µl | 1x |
| Vector | xµl | 20-100ng |
| Insert | xµl | 3:1 molar ratio over vector* |
| T4 DNA ligase 5U/µl | 2µl | 0.5U/µl |
* When restricted PCR products are ligated together, a 1:1 molar ratio is used. All the reagents are added following the order listed in the table above.
After finishing the reaction mixture, the content is mixed and incubated at room temperature (~22°C) for 20 min.
Reference: http://www.thermoscientificbio.com/uploadedFiles/Resources/fast-digestion-dna.pdf
iGEM 2013 Groningen
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