Key for Diagrams. “AND” Gate- both inputs are necessary for an output. Constitutive Promoter. Inducible Promoter. Repressible Promoter. Ribosome Binding Site. Open Reading Frame. Terminator. Retinoic Acid. BC-RA Converter Generator. Beta-Carotene. NO Input. Const. BC Generator.
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Key for Diagrams “AND” Gate- both inputs are necessary for an output. Constitutive Promoter Inducible Promoter Repressible Promoter Ribosome Binding Site Open Reading Frame Terminator
Retinoic Acid BC-RA Converter Generator Beta-Carotene NO Input Const. BC Generator Const. SoxR Generator 2 4 5 6 8 1 3 7 Metabolite Device 1- Oxidative Stress Induced RA Generator SoxS Promoter
Retinoic Acid NO SoxR BC-RA Converter NO Sensor Beta-Carotene Generator Device 1- Oxidative Stress Induced RA Generator SoxR gene SoxS Promoter Blh- BC to Retinal RALDH II- Retinal to RA Crt Cluster- BC Producer
Retinoic Acid NO SoxR BC to RA Converter NO Sensor B0015 TT k2230000 SoxR gene J23100 Const Prom. B0034 RBS ? Crt Cluster B0015 TT k2230001 SoxS Prom. k223035 Blh gene B0034 RBS B0015 TT k223036 Raldh II gene B0034 RBS J23100 Const Prom. B0034 RBS Beta-Carotene Generator Device 1- Oxidative Stress Induced RA Generator BC to Retinal Retinal to RA
Il-6 Secretion POPS Invertor 5 MT 5 4 3 2 1 Il-6 Generator Constitutive Export System Generator Device 2- Il-6 Secretion System
Il-6 Secreted Device 2 5MT Level Sensitive Sensor Il-6 Generator Repressible 5MT Operator Il-6 gene HlyA Linker Export System TolC gene Hly R,B,D,C
Il-6 Secreted Device 2 k223022 Il-6 + HlyA linker J23100 Const Prom. B0034 RBS k223025 Hly R,B,D,C B0015 TT k223024 TolC gene k223023 Trp Operator B0034 RBS B0015 TT 5MT Level Sensitive Sensor Il-6 Generator Allows Il-6 to leave through the export channel Export System Create an Export Channel and Chaperone
Device 2 Fourth Round of Cloning 2.4a Trp Operator, RBS,Il-6+HlyA,TT + Rbs, TrpR, TT (Cloned from step 2.3a) + (Cloned from step 2.1e) + J23100 + #? (Const. Promoter) (Synthesized de novo as 1part = RBS + trpR + TT) #k223023 + B0034 + k223022 + B0015 (Trp operator-) (RBS) (Il-6 and HlyAs linker) (TT) synthesized) Cut w/ EcoR1/Spe1 Cut w/ EcoR1/Xbal
Device 2 Fourth Round of Cloning 2.4b Trp Operator + Rbs, TolC, HlyR,B,D,C, TT (will be registered) + (Cloned from step 2.3b) + #k223023 (Trp operator - synthesized) B0034 + k223024 + k223025 + B0015 (RBS) (TolC ) (HlyR,B,D,C) (TT) Cut w/ ? Cut w/ ?