IGEM:Cambridge/2008/Notebook/Voltage/Mutant Strains

Mutant strains were required to identify and remove any interference from E. Coli's natural K+ homeostatic mechanisms. The following mutants were ordered from CGSC.

Strain Code	Mutation	Resistance
BW25113	Control	-
BW27564	Kdp E-F	-
BW30166	Kdp E-F	-
JW0045	Kef F	Kan
JW0683	Kdp D	Kan
JW1242	Kch	Kan
JW3314	Kef G	Kan
JW5096	Kdp E	Kan
TK3D11	Kdp	-

These strains were then plated out. The Kanamycin resistant strains were streaked onto kanamycin plates (50μg/mL) and also onto non-antibiotic plates. The non-resistant strains were plated onto plain plates. After growing overnight at 37°C single colonies were picked into 15μL broth (containing kanamycin if appropriate), and also streaked onto fresh plates. After growing overnight at 37°C glycerol stocks were made in triplicate and frozen at -20°C. The rest of the broth was stored at 4°C, as were the plates.

Checking Competence

To check all the mutants could be made competent All the mutant strains were then transformed with pUC19 and grown on ampicillin plates succesfully.