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Sensor: Week 6 June 20-25
From 2011.igem.org
Contents |
Tuesday June 21
Restriction Enzyme Test II
The sensor group digested the K3 vector with the following restriction digests:
| EcoRI | |
| EcoRI | XbaI |
| SpeI | PstI |
| EcoRI | PstI |
| XbaI | SpeI |
These digests were run on an agarose gel. The bands on the gel showed that the restriction enzymes were working.
Wednesday June 22
Radiation Sensor Construct, Day 1
The sensor group continued to assemble their test construct by placing a constitutive promoter in front of two RBS + cI repressor. The sensor group also attempted to place the OR site in front of the test construct pieces (K648000+K648001) and in front of an RBS and reporter. The sensor group tried to clone Cro into K3 and put an RBS in front of the cI repressor.
| Assembly Step | Part Involved with Assembly Step | |
|---|---|---|
| Insert PCR | J23113 B0034+mCherry K648000 | |
| Restriction Digest | Inserts using XbaI and PstI: | B0034+mCherry K648000+K648001 Cro C0051 |
| Insert using EcoRI and SpeI: | J23113 | |
| Vectors using SpeI and PstI: | K3+OR B0032 | |
| Vector using EcoRI and XbaI: | B0031+C0051 | |
| Vector using XbaI and PstI: | K3+RFP | |
| Ligation | OR + B0034+mCherry OR + K648000+K648001 J23113 + B0031+C0051 K3 + Cro B0032 + C0051 J23113 + B0030+C0051 | |
| Transformation/Plating | The above ligations and K3+OR were transformed into Escherichia coli cells. All cells were plated on plates with appropriate resistance. | |
B0034+C0051 and K3 were grown overnight in culture in order to extract more plasmid.
