Our project is starting to pick up! After our initial sampling/sequencing period, we realized that there is actual DNA we can work with from the tanks. This past week, we started our actual sample collecting from the tropical tank. We collected 3 sets of samples from the sediment, walls, and water. Throughout the week, we extracted the DNA and ran PCR on all 9 samples (plus one negative control). Today, we completed the gel electrophoresis and got some unpleasant results. Unfortunately, we couldn’t see the primer bands and the DNA bands didn’t show up like we thought they would. This means something went wrong in our PCR, but we don’t know if it was during PCR16SA or PCR16SB. Well, it’s back to the drawing board! Starting next week, we will be re-running the PCR on the 9 samples and possible collecting more samples from other tanks.
Although this week’s results were a bust, we know that there is definitely some DNA present that we can work with. I’m sure we’ll be finding some pretty cool things as we continue sampling and sequencing. 🙂
2 thoughts on “You win some, you lose some”
Hmm … usually when I have had PCR fail (which is alas not too rare) – I see big whopping primer bands (or, more like a smear at the bottom of the gel). Did you run a size marker lane (e..g, a DNA ladder)? Could you see DNA there? And what is PCR16SA vs PCR16SB?
We didn’t run a DNA ladder because we only had 10 wells and they were all used for the samples. Next week, we are definitely going to run the ladder with our samples. Those are the two programs we have programmed into our PCR machine. We run the first set of cycles, which we call 16SA, then we purify the samples and place it back into the machine for the second round of PCR, which is 16SB. We’re trying to be as organized as possible with our labeling to reduce the confusion which is bound to occur as the project progresses.