Monday 19^th September

Lab work

Visualization

Glycerol stocks of clones 7, 8, 9 and 10 containing FKBP - GFP 10 in pSB1C3

"By Maxence, Mahnaz Coline & Caroline"

The glycerol stock of the bacteria with the following plasmids were made.

• X (FKBP - GFP 10) clone 7
• X (FKBP - GFP 10) clone 8
• X (FKBP - GFP 10) clone 9
• X (FKBP - GFP 10) clone 10

500µL of liquid culture and 250 µL of glycerol 60% were mixed and put at -20°C.

Plasmids extraction of clones 7, 8, 9 and 10 containing GFP 1.9 in pSB1C3

"By Maxence, Mahnaz Coline & Caroline"

The following plasmids were extracted using the "Charge Switch-Pro Plasmid Miniprep" kit:

• X (FKBP - GFP 10) clone 7
• X (FKBP - GFP 10) clone 8
• X (FKBP - GFP 10) clone 9
• X (FKBP - GFP 10) clone 10

PCR of pSB1C3 from dCas9 ST - GFP 11 clone 8 to correct mutations

By Maxence, Mahnaz, Coline & Caroline

In order to correct mutations in dCas9 ST - GFP 11, PCR was performed with the following protocol.

For each 50μl of reaction, mix the following reagents :

• 1 µL of matrix
• 1 µL of dNTPs (10mM)
• 2.5 µL of each primer mix (10µM)
• 10 µL of buffer HF (5X)
• 0,5 µL of Phusion polymerase
• 32.5 µL of nuclease free water

Mix gently and aliquot 50 μl of the mix into the PCR tubes on ice. Perform PCR as follow:

Primers used were: