10/01/2014

Today we started the experiment for bioaccumulation. The main idea is to let the bacteria let bacteria E. coli DH5α with our construction (BB_Essential + K346004) in liquid medium and after, quantify how much mercury is on nutritive medium and how much it has inside the bacteria. This is due to the bacteria capacity to accumulate mercury inside it.

For that, it was tested four different concentrations (0.5 ug/ml; 0.25 ug/ml; 0.1ug/ml; 0.05ug/ml) besides of a sample without mercury. We used as control the same bacteria linage with a gene not related to mercury metabolism (GFP) to analyze the influence of the mercury binding proteins naturally expressed by bacteria.

The procedures were made based on bio detection’s procedures. We also continued the validation and characterization of our parts regarding to genetic-molecular level.

Backing to the rigorous digestions of our bio bricks with the following system for BB_Essential + E0840, BB_Essential + K346004 and BB_Essential + BB_Bioremediation:

Electrophoretic profile of enzymatic digestion BB_Essential + E0840:

1: BB_Essential + E0840 in PSB1C3 not digested

2: BB_Essential + E0840 in PSB1C3 digested with EcoRI

3: BB_Essential + E0840 in PSB1C3 digested with PstI

4: BB_Essential + E0840 in PSB1C3 digested with EcoRI e PstI

Electrophoretic profile of enzymatic digestion BB_Essential + K346004:

1: BB_Essential + K346004 in PSB1C3 not digested

2: BB_Essential + K346004 in PSB1C3 digested with EcoRI

3: BB_Essential + K346004 in PSB1C3 digested with PstI

4: BB_Essential + K346004 in PSB1C3 digested with EcoRI e PstI

Electrophoretic profile of enzymatic digestion BB_Essential + BB_Bioremediation:

1: BB_Essential + BB_Bioremediation in PSB1C3 not digested

2: BB_Essential + BB_Bioremediation in PSB1C3 digested with EcoRI

3: BB_Essential + BB_Bioremediation in PSB1C3 digested with PstI

4: BB_Essential + BB_Bioremediation in PSB1C3 digested with EcoRI e PstI

Regarding to the test of DH5α com BB_Essential + BB_Biooremediation that was made on September 30th, we didn’t get good results. We made again the inoculum in liquid LM with 20ug/ml, 50ug/ml, 70ug/ml and 100ug/ml of mercury chloride!