In which we calculate Molarity and get our First Results

The past few days have gone by quickly! Apparently, we did six days of work in our first five-day week, which was satisfying to learn. By the end of last week we had made our binding buffers, added them to our samples, and centrifuged them, and set up our Long Range PCRs.

However, Friday was the 13th, so some mishaps were to be expected. The Binding Buffer called for 39.8 g of Guanidine, a solid that looks like sugar that’s been sitting in someone’s china cabinet for 10 years. Guanidine becomes Guanidium in water, although it is notoriously hard to dissolve. Even still, 39.8 g seemed like too much, so we ran around and did math until we figured out that it was supposed to be 23.8825 g. Today, Heather found out that the protocol we had been using had miscalculations, and that we were correct to use 32.8825 g of Guanidine. That was a relief!

We have a vortex in the Ancient lab, but it strangely didn’t come with a plug. The cord just ended in a series of wires. Heather bought a plug and we wired it ourselves to no avail. In the end, we used our inner human vortexes and it was fine. At the same time, our pH meter didn’t want to calibrate, so Emily spent a long time fiddling with it. We needed to get the pH of the sodium acetate to 5.2. Eventually, we got close enough.

On Monday, we had a shorter day. In the morning, we made several Wash Buffers in the Ancient lab. Everything went smoothly. The only hiccup was when we discovered our SDS was 10% instead of 20%, so we compensated by using twice as much and adjusting the amount of water added to the solution. After lunch, we went up to the modern lab and made a gel to run the Long Range PCRs on. Previously, I had only used pre-cast gels, so it was satisfying to make the gel ourselves. We pipetted our DNA into the wells as quickly as possible so that the DNA couldn’t leak out. After the gel was finished running, we looked it on an imager. The gel (below) showed excellent amplification of partial mitochondrial genomes for the modern beaver!

LR PCR Gel 16 May 2016

Gel of Beaver Mitochondrial DNA: There are two rows. The two long smears on each end of the top row and in the second to last lane of the bottom row show a DNA ladder with known (short!) sizes of DNA fragment. The other bands on the top row are ~11,500 bps of DNA, while the bottom row has ~6,000bp fragments of DNA. 

Now we can go forward turning those sequences into baits to fish out ancient DNA.

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