Team:Exeter/Results

Green Light Module

We have had marked success with our Green Light Module. Not only is the magenta pigment bright and evenly synthesised, but we have also managed to show magenta synthesis is inhibited by exposure to white light. We are currently working on showing that green light specifically switches off magenta production.

Universal showing our vibrant magenta pigment

Plates grown in the dark clearly show magenta pigment

Comparison of liquid cultures grown in dark vs. light

Cyan pigment

We managed to get our cyan pigment working in the labs after some difficulties with our digestions/ligations. The pigment takes a while to develop, over 24 hours compared to magenta's overnight development, but the colour is reasonably strong and is produced evenly by the cells. However, more time has been invested in the cyan pigment which is responsive to red light (see "Planned Post-Freeze Work").

Red Light Module output Brick with BglII and BamHI restriction sites

Our output brick for the Red Light Module includes the ompF promoter which is specific to the protein [http://en.wikipedia.org/wiki/Porin_response:_osmoregulation_in_E.coli OmpR]. This is followed by an RBS and a restriction site for BglII and the coding region for a [http://parts.igem.org/Part:BBa_E0040 GFP]. A second restriction site for BamHI follows, then a series of STOP codons to act as a terminator. The whole sequence is flanked by the iGEM prefix and suffix allowing the BioBrick to be used in 3A Assembly.

We designed this BioBrick to be built by IDT in their gBlock format. The BioBrick would be formed from two gBlocks with the "scar" between them falling some way into the coding region of the GFP.

The two ends of GFP have been ligated together to form a complete, working protein in the successful Gibson assembly of our output bricks. GFP is constitutively expressed within DH5α cells as endogenous ENVZ phosporylates endogenous OmpR as a component of osmolarity sensing. Endogenous OmpR then binds to OmpF. Thus our output brick is working as expected. In order to test whether the output brick can be implemented into the Red light pathway and ENVZ- strain should be used to avoid false positives.

Planned "Post-Freeze" Work

Blue Light Module

Our Blue Light Module, which is being built by DNA2.0, will unfortunately not arrive in the labs until after the Wiki-freeze. However, we will be bringing plenty of pictures of it working (hopefully!) to the Jamboree. Considering it has been constructed in a very similar manner to the Green Light Module, which is functioning in the lab, we are reasonably hopefully that we can have the Blue Light Module functioning in the lab soon after its arrival.

Red Light Module

(Cyan working in EnvZ deficient cells)