So we’ve done our data collection, we’ve done more PCR than we could ever imagine, and we finally got our sequences. Now what? We analyze our data using Qiime, a software that will help us see connections between our microbe data. Qiime is useful since it can handle the large amounts of data we’re throwing at it, something most other programs would crash just thinking about. As someone with average knowledge of computers, it’s entirely intimidating learning something based on programming from scratch, but it has also been a great learning experience. Now off to play with terminal and find some meaning in these strings of letters…
Finally back in the lab
After spending the majority of our time collecting samples and doing water chemistry on site, we are all back in our lab ready to do DNA extractions and PCR galore. Unfortunately, the kim wipes we used to scrub microbes off the walls of the tanks are too large to fit into our tubes with the extraction beads. After about an hour of stressing and improvising, we were able to get a usable amount of supernatant…or so we hope. We will find out after we do our PCR.
Sample testing using new equipment
We finally got all our equipment for gathering metadata on the water, and decided to do some sample testing using tap water and saltwater from the tank. We ended up with the following:
Tap water –
NO2-N levels: 65 ppm
NO2 levels: 154.63 ppm
Phosphorus: 105 ppb
NO2-N levels: [TRIAL ONE] 65 ppm, [TRIAL TWO] 57 ppm
NO2 levels: [TRIAL ONE] 213.85 ppm, [TRIAL TWO] 187.53 ppm
Phosphorus: [TRIAL ONE] 48 ppb, [TRIAL TWO] 200 ppb
Some problems we encountered: When I did the trial one on the saltwater phosphorus test, I used the wrong reagent (phosphate reagent is used for the nitrite scanner, while phosphorus reagent is used for the phosphorus scanner). In addition, the phosphorus scanner maxes out at 200 (Andrew later confirmed this by doing additional scans on the saltwater), which means we’ll need a broader range scanner.
New Parameters to Test!
We’ve ordered and are recieving a series of equipment to measure Nitrate, Nitrite, Ammonia, dissolved Oxygen, pH, salinity, temperature, Phosphate, alkalinity, Chloride, hardness, Iron, and Sulfite in the tanks’ waters. Now we can gather more information about the environment these microbes are thriving in.
Sampling, DNA extraction, and PCR, OH MY!
For the past week, we’ve been doing preliminary sampling, as well as some DNA extractions and PCR on samples from the saltwater and freshwater flasks of water that we filtered and the wall and protein skimmer from the saltwater tank. Those four samples have now been through PCR. Although we found DNA, but the gel prepared after the PCR failed. We also went back to the tanks to gather samples from a saltwater tank that is soon to be broken up to create two coral ponds.
Yes, a boring title, but that’s exactly what this is. I am Alex Alexiev, second year biological sciences major. I’ve been thinking about a microbiology major for the past year or so, and this lab seems like a step in the right direction. I love aquatic systems and I think microbes are awesome, so I am extremely excited to research the two. Who knows what results the data will yield, but at the very least, it should be interesting.