Header

"Confocal Microscopy"
- Preparation of all the samples listed the previous day for Immunostaining with FITC conjugated antistreptavidin antibody.
- Preparation of all the samples listed the previous day for fluorescent staining with FITC conjugated biotin.
- Confocal afternoon.

Sensing-Effector

DpnI digest of the iGEM promoter and the GelE Backbone
-I digested the newly amplified and purified PCR products. The concentrations were not too good but I went ahead with the Gibson Assembly.

Miniprep of the Cad-transformants&PCR thereof
-I did the MIniprep of the innoculated cad Transformants and then I did a 20ul reaction PCR in order to check for the presence of the Cad promoter insert. The PCR showed that it was indeed present.

Gibson Assembly of the constitutive promoter+GFP (sensing) and the GelE+GFP construct (effector)
-I assembled the constitutive promoter in the plasmid in front of GFP so that we could use it as a positive control for the sensing module. I also inserted the GelE gelatinase gene into the plasmid containing the arabinose sensitve promoter.

Colony PCR of the MMP9 construct
-In order to check for the presence of the MMP9 insert I did a colony PCR with the colonies that were transformed with this plasmid. But the colony PCR did not work so I decided to innoculate the colonies and do the PCR on the MiniPrep the next day.

Transformation of cells with the constitutive promoter+GFP and the GelE gelatinase gene
-I transformed competent cells with the two last constructs, the one that would serve as a positive control for the sensing module (constitutive Promoter+GFP) and the other one with the gelatinase gene for GelE.

Note
I also realized that the constructs for the effector module would not express GFP because due to the primers there was a frame shift that induced a stop codon just after the linker and before GFP.

Nanoparticles

DLS measurements