Figure 3

Diagrams of the plasmids constructed to enable variation of genetic device copy number and to determine DTCs. A. Plasmid backbone with the nptII selection marker. The origins of the plasmids pSC101, p15A, pMB1, and pUC were cloned into the backbone using the two unique restriction sites SmaI and AvrII (not shown). TT denotes transcriptional terminator. Two different terminators (λ t0 and rrnB T1) had to be used because cloning attempts aimed at having the same terminator present simultaneously in opposing directions met with failure. The cassettes depicted in panels B and C were cloned into the multi-cloning site indicated by an asterisk. B. The cat device containing cassette cloned into the backbone. C. The cat and gfp device containing cassettes cloned into the backbone to arrive at the model system. Spacers were used to create spatial separation between neighboring devices (see Methods) to minimize the potential of spatial coupling. The promoter driving cat in B is different from that in C. While the native promoter was used in the former, P_L was used in the latter.