Goal: To persuade a bacterium to produce a variety of small peptides and secrete them into the medium

Standard BioBrick plasmid backbone Peptide secretor cassette origin antibiotic resistance X E PS (

Cassette Structure GFP Secretory signal RBS M Peptide His

Standard Promoter e.g. J23100 – Strong, constitutive promoter GFP Secretory signal RBS M Peptide His

Standard RBS e. g. B0034 – star 1 and W GFP Secretory signal RBS M Peptide His

Synthesize Secretory Signal e. g. lipA or sacB – identified from registry Possibly need to synthesize Leave as an exercise for the student! GFP Secretory signal RBS M Peptide His

Some random gene Need a signaller… … and a handle Allow student contributions for alternatives GFP Secretory signal RBS M Peptide His

Need: – 1 cleavage site – 1 target peptide – 1 purification handle The ‘slot’ in the cassette GFP Secretory signal RBS M Peptide His

What goes in the player? The polymer engineer is thinking about, e.g. extracellular matrix proto-molecules and self- assembling elements The biologist is thinking about using this as a recruiting tool… GFP Secretory signal RBS M Peptide His