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EPF-Lausanne/4 August 2009
From 2009.igem.org
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We ran an agarose gel as a control for PCR and digestion products from the previous day: LacI-LovTap-Term E/S, LacLP-LovTap-Term E/S, T7-LovTap-Term E/S, death cass. E/S, LovTap-TERM made by PCR only (new primers), PCR neg. control, PCR pos. control, LovTap-Term E/S, death cass. E/S (again). Everything seemed to go as planned =) | We ran an agarose gel as a control for PCR and digestion products from the previous day: LacI-LovTap-Term E/S, LacLP-LovTap-Term E/S, T7-LovTap-Term E/S, death cass. E/S, LovTap-TERM made by PCR only (new primers), PCR neg. control, PCR pos. control, LovTap-Term E/S, death cass. E/S (again). Everything seemed to go as planned =) | ||
| - | As the sequencing of our first LovTap-Term might not have worked well, we did a digestion assay, cutting the LovTap-Term plasmid with PstI (this should normally give 3 fragments, since there are 2 PstI restriction sites in LovTap). | + | As the sequencing of our first LovTap-Term might not have worked well, we did a digestion assay, cutting the LovTap-Term plasmid with PstI (this should normally give 3 fragments, since there are 2 PstI restriction sites in LovTap). --> the gel gave the expected bands! |
We prepared 14 LB-Agar plates with antibiotics ampicillin and kanamycin. | We prepared 14 LB-Agar plates with antibiotics ampicillin and kanamycin. | ||
Transformed DH5-alpha cells with all ligation products from the overnight reaction (16 in total): LovTap-Term (which we redid in case the first LovTap-Term isn't correct) with phosphatase treatment, LovTap-Term non-phosph.-treatment, LovTap-Term neg. control, LacI-RBS-LovTap-Term clone n°2 phosph., LacI-RBS-LovTap-Term clone n°5 phosph., LacI-RBS-LovTap-Term clone n°7 phosph., LacI-RBS-LovTap-Term clone n°2 non-phosph., LacI-RBS-LovTap-Term clone n°5 non-phosph., LacI-RBS-LovTap-Term clone n°7 non-phosph., LacI-RBS-LovTap-Term neg. control, LacI-Lov_Term (from semi-2-step PCR)- death cass. vect., LacLP-LovTap-Term-d.c. vect., T7-LovTap-Term-d.c.vect., LovTap-Term (from PCR) n°1, LovTap-Term (from PCR) n°2. | Transformed DH5-alpha cells with all ligation products from the overnight reaction (16 in total): LovTap-Term (which we redid in case the first LovTap-Term isn't correct) with phosphatase treatment, LovTap-Term non-phosph.-treatment, LovTap-Term neg. control, LacI-RBS-LovTap-Term clone n°2 phosph., LacI-RBS-LovTap-Term clone n°5 phosph., LacI-RBS-LovTap-Term clone n°7 phosph., LacI-RBS-LovTap-Term clone n°2 non-phosph., LacI-RBS-LovTap-Term clone n°5 non-phosph., LacI-RBS-LovTap-Term clone n°7 non-phosph., LacI-RBS-LovTap-Term neg. control, LacI-Lov_Term (from semi-2-step PCR)- death cass. vect., LacLP-LovTap-Term-d.c. vect., T7-LovTap-Term-d.c.vect., LovTap-Term (from PCR) n°1, LovTap-Term (from PCR) n°2. | ||
| + | |||
| + | The plates were left to incubate overnight at 37°C. | ||
==People in the lab== | ==People in the lab== | ||
Latest revision as of 15:44, 4 August 2009
Contents |
Wet Lab
Miniprep of cell cultures for LovTap-TERM, LovTap, B0015 (terminator) and P1010 (death cassette).
We ran an agarose gel as a control for PCR and digestion products from the previous day: LacI-LovTap-Term E/S, LacLP-LovTap-Term E/S, T7-LovTap-Term E/S, death cass. E/S, LovTap-TERM made by PCR only (new primers), PCR neg. control, PCR pos. control, LovTap-Term E/S, death cass. E/S (again). Everything seemed to go as planned =)
As the sequencing of our first LovTap-Term might not have worked well, we did a digestion assay, cutting the LovTap-Term plasmid with PstI (this should normally give 3 fragments, since there are 2 PstI restriction sites in LovTap). --> the gel gave the expected bands!
We prepared 14 LB-Agar plates with antibiotics ampicillin and kanamycin.
Transformed DH5-alpha cells with all ligation products from the overnight reaction (16 in total): LovTap-Term (which we redid in case the first LovTap-Term isn't correct) with phosphatase treatment, LovTap-Term non-phosph.-treatment, LovTap-Term neg. control, LacI-RBS-LovTap-Term clone n°2 phosph., LacI-RBS-LovTap-Term clone n°5 phosph., LacI-RBS-LovTap-Term clone n°7 phosph., LacI-RBS-LovTap-Term clone n°2 non-phosph., LacI-RBS-LovTap-Term clone n°5 non-phosph., LacI-RBS-LovTap-Term clone n°7 non-phosph., LacI-RBS-LovTap-Term neg. control, LacI-Lov_Term (from semi-2-step PCR)- death cass. vect., LacLP-LovTap-Term-d.c. vect., T7-LovTap-Term-d.c.vect., LovTap-Term (from PCR) n°1, LovTap-Term (from PCR) n°2.
The plates were left to incubate overnight at 37°C.
People in the lab
Nath, Basile, Gab, Christian, Nicolas
