Into the laboratory I

After a lot more theory and computing, we've started what's known in the business as wet lab work: chemicals and pipetting and Eppendorf tubes. We're hoping to put our new-found knowledge about turning DNA sequence data into phylogenetic trees to use. Day one of three was about extracting DNA from organisms.

First, I took the tiniest little palm-sized mortar and pestle and ground up the samples. These were two bits of mystery leaf (probably spinach and coriander) and two crickets. The leaves were very simple, but the crickets had to be disassembled.

Apologies to entomologists who enjoy anatomical correctness

A) One back leg set aside for another analysis next week
B) All other legs removed
C) Head removed (there's apparently something in the head which interferes with the chemistry of the DNA-replicating reaction)
D) Guts squeezed out and the outside of the abdomen discarded

It was a mercy that the crickets didn't smell too strong. But squeezing out the innards of the big one was utterly repulsive.

With each ground-up sample in its own little tube, I then let them soak in various chemicals to separate the DNA from the rest of the stuff. To then physically separate the different materials, the tubes were spun in a centrifuge. This little machine (about half the size of a microwave) contains something like a tiny roulette wheel, with slots for tubes around the outside. Their bottoms face outwards. As it spins (super fast; making use of centrifugal force), the heaviest material in the tubes is forced to the very bottom. The next-lightest material forms a distinct layer on top of that, then the next-lightest material, and so on. Here, the DNA was lightest, so I had a nice big layer of the liquid holding the DNA at the top.

This liquid was removed to its own new tube, and the rest discarded. There's a real skill to using the pipettes. They're very well crafted for moving tiny amounts of liquid around, but the liquid sure can move fast. Despite having some experience with them I did manage to spray myself with chloroform when my pipette unexpectedly squirted.

So, with a reasonably purified DNA suspension, the next step was to get rid of the suspension and replace it with another liquid. I honestly cannot remember why. But, the first one was a carcinogen and I've already proven liable to spray it over me.

I first added something which made the DNA gather together into a visible cloudy mass. Another centrifuge packed this mass down into what's referred to as a pellet, though in practice it's more of a tiny whiteish smear.

I sucked out all of the liquid I could and then rinsed my tiny whiteish smears in ethanol to dissolve anything else away. I did suck out most of the ethanol too, but because it evaporates so easily the final drying out was done by a much easier hot surface. Finally, I added the presumably non-carcinogenic solvent and watched my tiny whiteish smears dissolve away again.

The last task of the day was to take out a tiny tiny amount from each of my tiny tubes of tiny sample and put them through a machine called a NanoDrop. This uses lasers to quite literally see how much DNA there is and how much contaminant is left. It's not the most accurate, but gave all of my samples a tick of approval, which was a pleasant surprise. I've done this sort of procedure once before, back in the depths of undergrad, and few of us managed to get it to work at all. So, day one for me was a success.

Wet lab is turning out to be less frightening than I thought.