Bacterial Biosensors

Split Luciferase Complementation Assay

Advantages of luciferase for our project :

• Fast and reversible signal (quasi-immediate)
• Light excitation not needed (minimal autofluorescence) => simplification of the signal detection and processing mechanisms
• Signal highly correlated with concentration of substrate => very modulable
• Small concentrations of substrate are needed

Our constructs :
N terminal part of Luciferase fused to CheZ and C terminal part of Luciferase fused to CheY.

In absence of chemoattractant, CheZ and CheY are interacting and the two parts of the split reassemble to oxidize luciferin and thus luminescence is triggered. After addition of a chemoattractant (e.g. Glucose), CheZ and CheY don't interact anymore and luminescence is shut down.

We created two different constructs, one with split Firefly luciferase and one with split Renilla luciferase, to test different possibilities with different substrates (Firefly luciferase oxidizes d-luciferin and Renilla luciferase oxidizes coelenterazine)

Data For Our Favorite New Parts

Documented & submitted:

• BBa_K1486001 (Ara CpxR http://parts.igem.org/Part:BBa_K1486001 )
• BBa_K1486002 (Ara GFPCpxR http://parts.igem.org/Part:BBa_K1486002 )
• BBa_K1486005 (Ara CpxrGFP http://parts.igem.org/Part:BBa_K1486005 )
• BBa_K1486043 (LeuZ + rLuc http://parts.igem.org/Part:BBa_K1486043 )
• BBa_K1486049 (CpxR reporter promoter FW http://parts.igem.org/Part:BBa_K1486049 )
• BBa_K1486050 (CpxR reporter promoter RV http://parts.igem.org/Part:BBa_K1486050 )
• BBa_K1486056 (Ga1Mut http:/parts.igem.org/Part:BBa_K1486056 )
• BBa_K1486054 (CheY/CheZ + split rLuc http://parts.igem.org/Part:BBa_K1486054 )
• BBa_K1486055 (CheY/CheZ + split fLuc http://parts.igem.org/Part:BBa_K1486055 )

Works as expected:

• BBa_K1486002 (Ara GFPCpxR http://parts.igem.org/Part:BBa_K1486002 )
• BBa_K1486005 (Ara CpxRGFP http://parts.igem.org/Part:BBa_K1486005 )
• BBa_K1486008 (GA1 TedCécile http://parts.igem.org/Part:BBa_K1486008 )
• BBa_K1486018 (Ara split fLuc http://parts.igem.org/Part:BBa_K1486018 )
• BBa_K1486021 (Ara split rLuc http://parts.igem.org/Part:BBa_K1486021 )

Works as expected & submitted:

• BBa_K1486002 (Ara GFPCpxR http://parts.igem.org/Part:BBa_K1486002 )
• BBa_K1486005 (Ara CpxrGFP http://parts.igem.org/Part:BBa_K1486005 )
• BBa_K1486018 (Ara split fLuc http://parts.igem.org/Part:BBa_K1486018 )
• BBa_K1486021 (Ara split rLuc http://parts.igem.org/Part:BBa_K1486021 )
• BBa_K1486049 or BBaK1486050 (CpxR reporter promoter FW or RV http://parts.igem.org/Part:BBa_K1486050 )

Data For Pre-existing Parts

Documented & submitted: BBa_K1486056 (CpxR reporter (Calgary) http://parts.igem.org/Part:BBa_K1486056 ) We repaired it

We've Also Characterized the Following Parts