1. BioSketchHarvard iGEM 2005 1 BioBrick: Aggregate Parts Quad Part Inverter (QPI) RBS CodingRegion Terminator Promoter RBSTerminator ReporterCodingRegion Reporter

2. BioSketchHarvard iGEM 2005 2 QPIs and Reporters LacI QPI GFP (mut3b) Lambda cI QPI 434 cI QPI Have LacI(ts) QPI Venus Lambda cI857 QPI Need mCherry

3. BioSketchHarvard iGEM 2005 3 Test Constructs GFP (mut3b) LacI QPI GFP (mut3b) Lambda cI QPI GFP (mut3b) 434 QPI P lambda P Lac GFP (mut3b) LacI(ts) QPI GFP (mut3b) 857 QPI P 434 P lambda LacI QPI P lambda GFP (mut3b) Lambda cI QPI P lambda GFP (mut3b) Lambda cI QPI LacI(ts) QPI P 434 GFP (mut3b) 434 QPI 857 QPI

4. BioSketchHarvard iGEM 2005 4 BioBricks Testing Condition Temp (° C) 3030303040404040 IPTG(-)(-)(+)(+)(-)(-)(+)(+) UV(-)(+)(-)(+)(-)(+)(-)(+) QPIsLacI(-)(-)(+)(+)(-)(-)(+)(+) lambda(-)(+)(-)(+)(-)(+)(-)(+) 434(-)(+)(-)(+)(-)(+)(-)(+) 857(-)(-)(-)(-)(+)(+)(+)(+) LacI(ts)(-)(-)(+)(+)(+)(+)(+)(+) ConditionStimulusUVheatIPTGnone Initial State (-)(+)(-)(+)(-)(+)(-)(+) Toggle Switches LacI (Collins) (+)(+)(-)(+)(-)(-)(-)(+) 857(+)(+)(-)(-)(-)(+)(-)(+) LacI(ts)(+)(+)(-)(-)(-)(-)(-)(+)

5. BioSketchHarvard iGEM 2005 5 Things We Want to Know Minimal exposure time to UV to switch state Minimal concentration of IPTG to switch state Minimal exposure time to heat to switch state Amount of time a given state is maintained Time delay between input and reporting Effect of varying cell densities Effect of competing inputs Does SOS work in stationary phase Effectiveness of UV laser pointer

6. BioSketchHarvard iGEM 2005 6 Schedule - Week 1 Week 1MutationBegin introducing ts mutation into LacI coding region and LacI QPI BioBrick Assembly RBS + 857T + P lambda mCherry + TVenus + T P lambda + QPI LacI QPI lambda + GFP QPI 434 + GFP Collins

7. BioSketchHarvard iGEM 2005 7 Schedule - Week 2 Week 2MutationContinue mutations BioBrick Assembly RBS-857 + T-P lambda (QPI 857 ) P lambda -QPI Lac + QPI lambda -GFP P lambda -QPI Lac + GFP P lac + QPI lambda -GFP P lambda + QPI 434 -GFP RBS + mCherry-TRBS + Venus-T Collins

8. BioSketchHarvard iGEM 2005 8 Schedule - Week 3 Week 3MutationSequence mutations BioBrick Testing P lambda -QPI Lac -GFP P lac -QPI lambda -GFP P lambda -QPI 434 -GFP P lambda -QPI Lac -QPI lambda -GFP (Collins) BioBrick Assembly P 434 + QPI 857 Collins

9. BioSketchHarvard iGEM 2005 9 Schedule - Week 4 Week 4BioBrick Testing II P lambda -QPI Lac -GFP P lac -QPI lambda -GFP P lambda -QPI 434 -GFP P lambda -QPI Lac -QPI lambda -GFP (Collins) BioBrick Assembly P 434 -QPI 857 + GFP P 434 -QPI 857 + QPI 434 -GFP P lambda + QPI Lac(ts) Collins

10. BioSketchHarvard iGEM 2005 10 Schedule - Week 5 Week 5BioBrick Testing P 434 -QPI 857 -GFP P 434 -QPI 857 -QPI 434 -GFP (857 system) BioBrick Assembly P lambda -QPI Lac(ts) + GFP P lambda -QPI Lac(ts) + QPI lambda -GFP Collins

11. BioSketchHarvard iGEM 2005 11 Schedule - Week 6 Week 6BioBrick Testing II P 434 -QPI 857 -GFP P 434 -QPI 857 -QPI 434 -GFP (857 system) BioBrick Testing P lambda -QPI Lac(ts) -GFP P lambda -QPI Lac(ts) -QPI lambda -GFP (Lac(ts) system) Collins

12. BioSketchHarvard iGEM 2005 12 Schedule - Week 7+ Padding