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User:University of Washington/1 August 2008
From 2008.igem.org
LuxR from AraC and TetR
- BBa_P1010 (contains death gene) grew in both XL1-Blue and DB3.1 which was weird. XL1-Blue didn't have death resistance, so the cell shouldn't grow. Will miniprep and sequence the part P1010. But will use the vector plasmid containing Amp resistance gene from the part nevertheless.
- Obtained P1010 on pSB3K3 and pSB1AC3 from Brandi by inoculated the strain on selective plates.
Cure MG1655Z1
- Restriction digested the Elowitz's plasmid from original plate and stock plate with XbaI and BamHI (both cut both plasmid pACYC184 and pCD26 once)
| Reagents | Ori + XbaI | Stock + XbaI | Ori + BamHI | Stock + BamHI | Control |
|---|---|---|---|---|---|
| dH2O | 34 ul | 34 ul | 39 ul | 39 ul | 39.5 ul |
| Buffer | 5 ul NEB2 | 5 ul NEB2 | 5 ul NEB3 | 5 ul NEB3 | 5 ul NEB3 |
| BSA | 0.5 ul | 0.5 ul | 0.5 ul | 0.5 ul | 0.5 ul |
| DNA | 10 ul | 10 ul | 5 ul | 5 ul | 5 ul |
| vortex | |||||
| Enzyme | 0.5 ul XbaI | 0.5 ul XbaI | 0.5 ul BamHI | 0.5 ul BamHI | - |
| centrifuge and incubate 37 degree Celsius | |||||
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