By guest writer and DOP volunteer, Dr. Helen Ougham
In my previous blog, I explained how buccal swabs had been taken from the insides of the osprey chicks' cheeks at ringing time and stored in small, capped plastic tubes in a fridge until the scientists were ready to use them. What happens next?
Aeron (and his two sisters, Menai and Eitha) kindly donates some of their DNA in the summer
Aeron. © MWT
Well, the buccal swabs will be a mixture of osprey cheek cells and osprey saliva, probably with bits of their last meal, and other things that they took in from their mum and dad during feeding - grass? ants? who knows...? What we really want is the DNA out of it.
An osprey's saliva sample will contain more than just osprey DNA!
Aeron, Menai, Glesni, Eitha. © MWT
In principle it's not that difficult to extract DNA from living cells. If you'd like to try it as a fun experiment with your children or grandchildren, the easiest way is to use fruit like kiwi fruit, bananas or strawberries, though it also works well with leafy vegetables, dried peas or beans and meat like liver. Apart from some ordinary kitchen equipment, all you'll need is washing-up liquid, salt and some alcohol -either methylated spirits or isopropyl alcohol ("rubbing alcohol") is the easiest to get hold of. There are many websites that will tell you how to do it; this one from the University of York is good, with plenty of illustrations.
At the end of an experiment like this, you and the kids will have some white, stringy-looking material - and that's the DNA from the fruit or vegetable you've used.
Here's a photo of DNA from the University of Utah's website on DNA extraction from split peas, another nice site.
The methods that work for fruit, split peas, liver and so forth would also work for osprey cells... but there's a snag. Each buccal swab only has quite a small sample of osprey cheek cells on it. How do we scale down the extraction process to get the DNA out of our swabs?
The answer to that is to use kits that have been specially developed for handling small samples. Nowadays there are quite a few companies that produce these. One of the best known is Qiagen (I've never been quite sure how to pronounce it).
They make a range of kits designed to give the best results with different kinds of samples - blood, plant material, bacteria and so forth. They even do one for getting DNA out of poo (or "DNA stool mini kit" as they more politely put it). Chris Packham from Springwatch would probably love this one!
It's kits from this company that are used in the labs at Aberystwyth. Each kit provides you with a set of small bottles of solutions and some special plastic tubes in which the DNA can be extracted from the cells and cleaned up - here's an example of what one of these kits contains.
A QIAamp DNA Mini Kit
In a couple of weeks' time, I'm going to be going back into the lab myself, for the first time in quite a few years, to do some osprey DNA extraction, and I'm quite excited! Fortunately I'll be supervised by Ilze, a talented scientist who worked with Matt Hegarty on a red kite DNA project and has plenty of recent experience getting bird DNA out of different kinds of samples. Many thanks to Matt and Ilze for letting me take part in the work.
Ilze Skujina - you can follow her on Twitter here
One of the techniques that Ilze developed was a way of getting DNA out of feathers. That's a bigger challenge than working with buccal swabs, because there are fewer cells in a feather and they're harder to get at, but with Ilze's method we should be able to extract DNA from many of the feathers that have been collected at the different Welsh sites.
Some of these feathers are from the parent birds and that will add a lot of useful genetic information to what we learn from the chick buccal swabs. Of course, we can't take swabs from the adult birds because they don't have the thanatosis ("playing dead") response that chicks do; it's thanatosis that makes it safe - for both the chicks and the ringers - to do the ringing and take the swabs at the same time. So feathers are a useful, if challenging, alternative.
Clarach and Cerist beautifully demonstrating their thanatosis technique back in 2013
Once we've extracted the DNA from our osprey samples, there are still a couple of difficulties. Firstly, the amount of DNA will be very small - not enough to let us do all the studies we want. Secondly, remember that at the start of this blog I mentioned that the swabs won't just have osprey cheek cells on them? They may also have fish from the chick's last meal, bits of plant material, maybe insects; and there'll be bacteria that live inside the osprey's mouth. All of these things have DNA of their own. How do we make sure that what we're looking at is just osprey DNA and osprey genes?
The answer to both questions is a very clever lab method that was first developed back in the 1980s. It's used a lot in forensic science, and much more widely in medicine and biology, and it's vital for the sort of osprey genetics research we're interested in. It's known as PCR for short (Polymerase chain reaction), and in the next blog I'll tell you about how it works and what we use it for with our osprey samples.
Meanwhile, wish me luck in the lab!
