Biological Equivalence Problem
Dealing with biological equivalence
Mathematically, the permutations (1,2) and (-2,-1) are not hard to distinguish. However, in a biological system, these two are indistinguishable. Transcription can occur in either direction (either the plus strand or minus strand) as long as the promoter is facing towards the coding region. A promoter followed by RBS-Tet (1,2) and a reversed RBS-Tet followed by a reversed promoter (-2,-1) will both be expressed (tetracycline resistant). Colonies carrying either of these configurations will grow on media containing tetracycline (as illustrated below). Thus, (1,2) and (-2,-1) are biologically equivalent.
To be able to distinguish the two permutations, we generated a reverse RFP reporter (BBa_J31011) to be placed as a stationary unit to the left of the pancake stack. In any configuration of pancakes where the promoter is in the reverse orientation, the cells will express RFP and glow red.