EPF-Lausanne/2 September 2009

Wet Lab

• Results of the culture

The RFP for Miniprep grew normally

The "positive control" of RO2 #4, #5, #10 on LB grew without showing redish color

All the other medium did not grow except for clone #5 in M9/min + AA+ thiamine, which shows litte turbidity

-> we let them a little bit longer in the incubator

• Glycerol stock

Clones #1 and #2 of CFP and RFP were made.

• Miniprep of the overnight culture of RFP (I13507) was minipreped.

• Digestion -> the host vector for the TrpO containing RFP (I13507) was digested. Enzymes : EcoRI-HF / XbaI.

• Results : the overnight ligation (started Monday) and transformed + incubated yesterday didn't give any results. None of the plates grew.

• Induction tests : the two overnight cultures of CFP have been induced at 83 ng/ml of ATC. The two overnight cultures of RFP (LacI inducible) have been induced at 1 mM of IPTG.

• Gel : to run the digested RFP and the TrpO.

• Ligation -> cloning the RO1. We used 20 ng of vector RFP (instead of 10). We did 3 different molar ratio for the insert : 6:1, 10:1, 12:1.

• Transformation : the 3 ligation products have been transformed according to the iGEM protocol.

• Results from the induction : the LacI inducible RFP turned red as expected (clearly secable after pellet was formed). The CFP TetR inducible didn't change colour. But it was slightly greenish... further experiments will be needed to check its functionnality and make sure there is no TetR gene in DH5a.

People in the lab

Basile, Mélanie, Caroline