Team:University College London/Protocols/CharacterisationSalt

Salt Tolerance

Step 1 - Setting up Falcons: Divide the prepared media into autoclaved shaker flasks such that you have two flasks containing equal amounts of each media

Step 2 - Inoculate: Inoculate one of each pair of flasks with 50ul of untransformed E.coli W3110 and the other with 50ul K398108 transformed cells.

Step 3 - Sampling: Mix and take 1ml sample from each flask – return the flask to shaker. Take Optical Density readings of each sample – record time of inoculation and Optical Density in a spreadsheet. (In order to allow for the time taken measuring Optical Density, the inoculations for each type of media should be staggered by approx. 10-15 mins). Sample each shake flask at approx one hour intervals from inoculation

NB: after a few readings dilutions will be required as the OD reading is not reliable if higher than 0.4 – ensure that readings are then multiplied by the dilution factor before recording in spreadsheet

Step 4 - Stopping protocol: Stop once all growth curves have come out of exponential phase (or it’s ridiculously late and sleep is required)