Figure 1.

Schematic representation of the chromosome from the two E. coli strains used here. ENS32 harbors an untranslated version of the lacZ gene (lacZΔ), and ENS304 harbors a gene encoding RNAI, a down-regulator of ColEI plasmid replication. The black box represents the T7 gene1, encoding T7 RNAP. P_T7 corresponds to the T7 gene10 promoter from positions −21 to +2 with respect to the natural transcription start; Op (operator) represents the first 22 transcribed nucleotides from the lac operon encompassing the high-affinity lac repressor-binding site. Ter (terminator) represents the T7 late transcription terminator (Tφ), fused to the trp rho-independent terminator in the case of ENS304 (cf. 19). In ENS32, the operator is followed by the short polylinker from plasmid pEMBLΔ46 (46) and then by a truncated version of the lac operon extending from nucleotides +58 to 4010 with respect to the genuine transcriptional start, i.e., lacking the lacZ ribosome-binding site. In ENS304, the operator is followed by a BamHI linker and then by the genuine pBR322 RNAI gene (Fig. 3a). In both strains, the endogenous lacZ gene has been inactivated by Tn10 insertion (ENS32), followed by imperfect excision (ENS304).