Monday 12^th July

Lab work

Getting DNA closer

Amplification of DS-TDcasN-, DS-SPcasN- and pZA21 plasmids

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^® High-Fidelity 2X Master Mix from usual protocol adapted to get 50µL at the end.

Biobrick characterization

Electrocompetents and transformation by electroporation of BL21

By Charlene

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. 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)
• 30µL of bacteria + 3mL LB + Chloramphenicol (30µg/mL)

Cells were incubated for 8h at 37°C, 200 RPM.

No bacteria grew so we concluded that there was a problem with the transformation.