Difference between revisions of "Team:Paris Saclay/Notebook/July/12"
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==Lab work== | ==Lab work== | ||
| − | === | + | ===Getting DNA closer=== |
====Amplification of DS-TDcasN-, DS-SPcasN- and pZA21 plasmids==== | ====Amplification of DS-TDcasN-, DS-SPcasN- and pZA21 plasmids==== | ||
''By Caroline'' | ''By Caroline'' | ||
| − | Plasmids extracted the 4/07/16 were used to amplify the dCas9 needed for the get DNA closer tool and the pZA21 plasmid in which the tool will be inserted. A PCR was carry out following PCR with Q5<sup>®</sup> High-Fidelity 2X Master Mix from | + | Plasmids extracted the 4/07/16 were used to amplify the dCas9 needed for the get DNA closer tool and the pZA21 plasmid in which the tool will be inserted. A PCR was carry out following PCR with Q5<sup>®</sup> High-Fidelity 2X Master Mix from NEB protocol (cf protocols section) adapted to have 50µL at the end. |
| − | === | + | ===Electrophoresis of the PCR amplification=== |
| − | ''By | + | ''By Caroline'' |
| − | + | 5µL of the amplifications are mixed with 1µL of purple loading dye. The solutions are put into a 0.8% agarose gel and migrated for 25min. | |
| − | + | ||
===Biobrick characterization=== | ===Biobrick characterization=== | ||
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Cells transformed on 11/07/16 grew overnight. However no colony was observed. | Cells transformed on 11/07/16 grew overnight. However no colony was observed. | ||
| − | To control if there is a bacteria transformed which was not put on Petri dish, we performed another experiment. | + | To control if there is a bacteria transformed which was not put on Petri dish, we performed another experiment. 100 µL (500µL concentrated) of transformed bacteria for each condition, conserved at -4°C from yesterday, were split in 3 solutions : |
* 30µL of bacteria + 3mL LB + Streptomycin (50µg/mL) + Chloramphenicol (30µg/mL) | * 30µL of bacteria + 3mL LB + Streptomycin (50µg/mL) + Chloramphenicol (30µg/mL) | ||
* 30µL of bacteria + 3mL LB + Streptomycin (50µg/mL) | * 30µL of bacteria + 3mL LB + Streptomycin (50µg/mL) | ||
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No bacteria grew so we concluded that there was a problem with the transformation. | No bacteria grew so we concluded that there was a problem with the transformation. | ||
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{{Team:Paris_Saclay/notebook_footer}} | {{Team:Paris_Saclay/notebook_footer}} | ||
Revision as of 15:38, 12 July 2016
