pVIB onto E. coli - bacterial transformation (Sep/21/2005 )
So i am using pVIB, from Vibiro fischeri (an ampicillinent resistant, bioluminescent plasmid) and transferring it to E. coli. my experiemnt is what factors will increase or decrease the number of mutated E. coli colonies, allowing them to glow/ disallowing them to glow.
So far i am adding ampicillin, salinity, pH, and lactoferrin to the agar in increments of low , standard, and high (which may be over load, considering i dont want to kill the bacteria). I am also using temperature of incubation as a factor including a low, standard (room temp), and high. I am using a statistical analysis called Design of Experiment, which allows for no control in an experiment.
I am pre-testing this week but i would like to know if someone has ever tried adding stuff to the agar, or if it is even viable for my type of experiment. Any comments would be greatly appreciated!
You should be aware that the V. fischeri luminescence system will not function at 37C. To see light, you must grow the cells at 28C. There are other systems, such as the one from Photorhabdus luminescens that will function at 37C. You can grow the cells at 37C then transfer them to 28C for a few hours to observe luminescence.
yes im aware, my research says that they will luminescence at 30C or lower .