Fig. 5

The test of MtlR box and cre in vivo and in vitro. A The state of genetic switch based on MtlR box and cre site. State OFF: MtlR box and cre site were added to the genetic circuit, and no substrate added. State OFF: MtlR box and cre site were added to the genetic circuit, and glucose added. State ON: MtlR box was added to the genetic circuit, and inducers added. State ON: MtlR box and cre site were added to the genetic circuit, and inducers added. B The construction of genetic circuit based on MtlR box and cre site. Three cre motifs is ‘TGWNANCGNTNWCA’, ‘AGCTTT-Yx-AAAGCT’, ‘AAAGCT-Yx- AGCTTT’ (in which N represents any base and W represents A or T, in which N represents any base and W represents A or T). Three cre sites were designed based on their motifs, in which cre1 sequence was ‘TGACAGCGCTGTCA’, cre2 sequence was ‘AGCTTTATAAAGCT’, cre3 sequence was ‘AAAGCTATAGCTTT’. Three cre sites were inserted to the artificial promoter with different location, generating six promoters ‘A-4a, A-4b, A-4c, A-4d, A-4e, A-4f’. Six fragments (A-4a, A-4b, A-4c, A-4d, A-4e, A-4f) were selected as probes, EMSA was performed by CcpA (0, 1.2, 1.6 µM) for six fragments. C The ON/OFF state values of the genetic circuit. The OFF-state values of six promoters ‘A-4a, A-4b, A-4c, A-4d, A-4e, A-4f’ when no substrate added. The OFF-state values of six promoters ‘A-4a, A-4b, A-4c, A-4d, A-4e, A-4f’ when glucose added. The ON-state values of six promoters ‘A-4a, A-4b, A-4c, A-4d, A-4e, A-4f’ when mannitol added