This year, we're focusing on the LacI, TetR, and cI Lambda repressors and their respective promoters. Our goal is to obtain a variety of behaviors from each of these parts by changing their sequences randomly, or mutating them. With this range of behaviors, we'll be able to design systems to exhibit targeted behavior depending on the chosen repressor. This will be a valuable contribution to the registry which contains many parts that have a limited range of activities such as always strongly on or inducible.

Having a well-characterized range of activity of a single part will provide more opportunities to fine tune a system with an added layer or regulation. When coupled with another part that has a similar range of activity, a repressible promoter with a repressor for example, there is a combinatorial effect which gives the user even more control.

General Construct

We designed this construct for characterizing promoter mutants. When pBAD is induced with arabinose, the repressor of choice is transcribed leading to decreased levels of the reporter, GFP. The specific order in which the parts are depicted allows the user to swap in any promoter/repressor or promoter/activator pair using our regulatory characterization plasmid.

The mutant repressor characterization construct is identical to the promoter mutant construct with the exception that the user must put the wildtype promoter on the 5' end and a mutant repressor/activator on the 3' end.