Team:BYU Provo/Notebook/Cholera - Enzyme/October/Period1/Dailylog

10/1/13 - 10/3/13

We spent these days finalizing our presentation and poster with the data that we had gathered, and left for the North American Jamboree.

10/4/13 - 10/6/13

The North American iGem Jamboree! We had a great time, learned a lot from the other teams, and are excited to be moving on to the World Championships! Now it's time to get back to work and get more data.

10/9/13

We made a new 500 mL stock of our SSW-LB following the recipe listed on the protocols page. We also set up overnights of PIG92 in LB+AMP, so that it can be purified and placed in our parts registry. We also started new cholera overnights from our plated cholera.

10/10/13

We checked the PIG92 overnight that was started yesterday, but it was still clear. We started three more overnights of PIG92 in LB, LB+AMP, and LB+CAM. We will check tomorrow to see which of these is growing correctly.

10/11/13

The overnight of PIG92 that was started on 10/9 is nice and cloudy today, indicating growth of our bacteria. The overnights started yesterday are still clear, including the overnight in LB, so the bacteria just takes a little more time to grow up than we were expecting. We pelleted the overnight from 10/9 and placed it in the freezer for tomorrow.

Our cholera overnights don't show any growth yet, but as we are starting these overnights from plated cholera, it will take almost a week before we have good biofilm growth.

The SSW-LB that we prepared on 10/9 doesn't look right. It is darker than it should be and is really cloudy. We discarded this stock and made a new 500 mL stock following the recipe listed in the protocols section. The new stock looks right and is free of contamination.

Our DspB colony plate grew well, so we took eight individual colonies and ran Colony PCR on them following the protocol listed on the protocols page. The plate with the streaked individual colonies was placed in the 37° incubator overnight and the PCR was left running overnight.

10/12/13

We ran the plasmid prep kit on the pelleted PIG92 overnight. The purified plasmid was stored in the freezer in our parts registry.

We also ran our colony PCR products out on gel to check for the presence of our plasmid. We ran 3 uL of product with 2 uL of loading dye for each sample. It appeared that there were really faint bands on three of our eight samples, but the bands were so faint we couldn't tell if the plasmid was there or not. We also pulled the 8-colony DspB plate from the 37° incubator and stored it in the fridge until we can verify if any of the eight colonies contain our plasmid.

10/14/13

We re-ran our colony PCR products out on gel, but ran 5 uL of product rather than the 3 uL that we ran before. We were hoping that running more product would cause any bands to be more visible and allow us to detect them, but we got similar results. We decided to pick eight more colonies from the plate, ran colony PCR on them, and streaked them onto a LB-Amp plate and incubated it at 37°.

10/15/13

We ran a gel for the colony PCR products we started last night and colony 5 showed a bright band right around 1100 bp's which is the right size for DspB. We then started a 5 ml overnight in LB-Amp from its corresponding streak on the plate.