Selection, design and optimization of color expression cassettes

Selection and Design

A full color spectrum may be made by mixing three saturated colors, each reflecting light appropriate for the color sensors in the human eye. For the subtractive color relevant for mixes of bacterial color on a plate, the idea colors are cyan, yellow, and magenta. From part sequences in the Registry, we selected the the blue chromoprotein amiCFP (K592010), the yellow chromoprotein amilGFP (K592009), and the classic red fluorescent protein (RFP, E1010). These parts were selected due to their excellent presentation by the 2011 Uppsala iGEM team. We altered the sequences to remove KpnI sites, for convenience during cloning and assembly, designed custom ribosomal binding sites for each open reading frame using the Salis RBS calculator ( to give consistent medium-high expression levels (TIR: 50k), ordered the sequences synthesized as gBlocks (IDT), and assembled them. Promoters were selected from the Anderson collection of constitutive sigma 70 promoters of various strengths, with changes to xxx.

Unfortunately, initial assemblies did not produce color. We therefore undertook an optimization program, xxx, change of base strain (TG1 produces larger, more saturated colonies than TOP10, which we interpret as a consequence of more robust growth).

Experiment Result

After optimization, xxx

The resulting color cassettes have been submitted as xxx [photo of color plates]

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