Together with some colleagues from CCW, I recently had the opportunity to travel to Ireland to undertake some very important studies. Namely, how to extract Mammal DNA from hair, droppings and barn owl pellets!
The work was part of the Mammals in a Sustainable Environment (MISE) project, a partnership between Wales and Ireland, part funded by the European Regional Development Fund (ERDF).
The project is working with volunteers to survey and monitor mammal species of special conservation interest and aims to raise awareness and engage the public whilst learning more about our native mammals on both sides of the Irish Sea.
An Irish Pine marten demonstrates how hair tubes work. Photo Terry Whittaker
Target species include otter, pine marten, red squirrel, polecat, stoat, weasel, harvest mouse, dormouse and several bat species.
ID’ing our critters
The project lead partner, Waterford Institute of Technology (WIT), are working on new DNA techniques to enable us to identify mammal species, and in some cases identify their sex and genetic haplotype from non invasive samples such as hair and faeces.
A good sample - but it is possible to extract enough DNA from a single hair.
This will enable us to monitor species that are difficult (or impossible) to survey using other methods.
We have been collecting samples and sending them to WIT for analysis. Throughout our visit to the Emerald Isle we got a chance to see just what we had been putting our colleagues through!
To extract our DNA, we of course require capturing some samples first, and this is done via a non-invasive means. Hair is collected using hair tubes – whereby sticky pads are placed inside a baited tube with the hope that visiting mammals leave a few hairs behind when they come to take the bait.
Pine marten faecal samples ready for testing. It really is a glamorous job!
Over in Wales we have been using this technique to look for signs of Red squirrels in Mid-Wales and Snowdonia. In Waterford, our colleagues have a healthy population of Pine marten to survey, and this provided us with some hair and animal faeces samples to test.
Extracting the DNA
The first step was to mix our samples with a buffer. The bigger sample the better, but it is possible to extract enough DNA from a single hair.
Indeed, we learned just how difficult it was to prise a few precious hairs from a sticky pad – and this is just the beginning of a very tricky process.
We then put the samples through a series of steps – pipetting into a tube, filtering it through a buffer in a centrifuge before pipetting into another tube and spinning and filtering again.
Our inital sample is first pipetted into a tube.
The risk of cross contamination meant that we had to be extra careful, ensuring that we used a different pipette tip for each and every step – ever vigilant for stray droplets of DNA escaping.
We wouldn’t want to cause unnecessary excitement by finding Irish Pine marten DNA in a Welsh sample!
Eventually we were left with a tiny sample of solution containing our precious DNA – plenty to run all the tests we need [so we were assured!]
Barn owls are great lab assistants
Though we tested faeces and hair samples, we also tested Barn owl pellets. These were particularly interesting to me as a colleague from Anglesey had provided them, in the hope of finding Harvest mouse DNA.
Harvest mice are under-recorded in Wales, and one of the aims of the MISE project is to carry out surveys to try and learn more about their distribution.
Eventually, after much filtering and centrifuging, we are left with a tiny sample of solution containing our precious DNA.
We can do this through nest searches, but we consider local Barn owls are probably better at finding Harvest mice than we are!
By collecting Barn owl pellets from areas where we think Harvest mice can be found, these predators help us understand the distribution of the Harvest mouse.
Time for some results
Soon it was time for some answers, and this is where the real magic happens.
Once we had extracted our DNA from our hair, scats and pellets we were ready to run them through the Realtime PCR test to identify species.
The Realtime PCR is used to amplify and simultaneously quantify a targeted DNA molecule in our samples. In simple terms, we can use pre-prepared primers containing DNA from our target species, and look for a reaction with our samples.
The strength of the reaction tells us whether our samples contain the same strand of DNA as those in our target species.
After a few hours in the machine, we would find out what species our samples came from (or in the case of my Barn owl pellets, what species the barn owl had been eating!) The following morning, our hard earned results were ready.
MISE Project officer Andrew Harrington setting up the samples to run through the PCR machine.
The first discovery was that this genetics game was a lot harder than it looked!
Quite a few of our samples were contaminated, while everyone compared results to see who was guilty of having a shaky pippetting technique!
We did have some results though (phew!) . There was no real surprise to find that our very hairy sticky pads and scats were from Pine marten.
Our colleagues at WIT have been studying the local Pine marten population for some time, and have developed their genetics techniques so that they are now able to identify individual pine martens from their hair and scats.
The barn owl pellets were less successful. While there was evidence of Wood mice and Voles, no Harvest mouse DNA was apparent. I’ll have to keep searching for those elusive little mice!
We left with a better understanding of how tricky DNA extraction is and just what sort of information we can get from our samples. Now I need to keep those samples rolling in from North Wales to keep our Irish colleagues busy.
Watch this space for news of how you can help in the next hunt for elusive Welsh mammal DNA!
