IGEM:Groningen/Notebook/iGEM 2011/2011/05/30

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30-5-11



Clone promotors upstream RBS-GFP-DT for eventual measurements of the promotor activities.
Start working on strategy
The following promotors are located on the following vectors:
PBADaraC: pSB2K2
PcI: pSB2K2
PlasB: pSB1A1

Digest the promotor fragment with: EcoRI and SpeI
Digest the vector with EcoRI and XbaI
Cutmix:
8μl buffer
2μl EcoRI
2μl XbaI
8μl MQ water
so 10μl mix with 10μl sample

CutmixII:
8μl buffer
2μl EcoRI
2μl SpeI
8μl MQ water
so 10μl mix with 10μl sample

Ligation overnight, per sample:
1μl T4 DNA ligase
2μl T4 DNA ligase buffer
7μl insert
3μl plasmid
7μl MQ water