Cloned constructs

For the final Colisweeper circuit we plan a four plasmid system. The mine cells constitutively express LuxI and NagZ. In the Non-Mine cells LuxR and PhoA are expressed constitutively whereas Aes and GusA are expressed from pLux promoters with different sensitivities. To get to this system we tested different versions of the circuit with GFP. In the following table we list all the biobricks we used, the plasmids we cloned and what experiments we used them for. In general we used standard biobrick cloning techniques which you can find in the Methods section. Whenever we used PCR gene amplification for cloning, we list the primers used in the following table. To be able to co-transform different plasmids we used backbones with compatible origins of replication and resistance genes. In the table you can find which versions we used for which constructs.

Figure 1. Plasmids in mine and non-mine cells

GFP constructs
	Description	Cloning	Maps
1	Receiver cell construct for GFP diffusion experiments	BBa_J09855 backbone (SpeI, PstI) and BBa_E0840 insert (XbaI, PstI)
2	Library of the Receiver cell constructs	Using the BBa_J09855-E0840 construct a library with mutated pLux promoters was created through site-saturation mutagenesis to screen for promoters with changed sensitivities for OHHL:
3	Receiver cell construct for GFP experiments without the LuxR generating part	BBa_R0062 backbone (SpeI, PstI) and BBa_E0840 insert (XbaI, PstI)
4			...

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