I Will Always Love Ewe - the story of Dolly the Sheep
Dolly the Sheep via Flickr, Toni Barros (CC BY-SA 2.0)
Cloning is a popular trope in science fiction, from Kazuo Ishiguro’s moving human cloning novel Never Let Me Go to Jurassic Park and that really bad Star Wars movie.
We’ll be taking a look at the science of de-extinction a la Jurassic Park in a future episode, and raising an army of aliens is out of scope for this podcast.
But cloning humans took a step from the realms of sci-fi towards reality in 1996, with the birth of Dolly the Sheep, the first live-born adult mammal clone created by taking the DNA from an adult cell and putting it into an egg.
Dolly’s birth was announced in February 1997, although she was actually born a few months earlier in July 1996. I was at university when I first saw the now infamous issue of the scientific journal Nature sporting a rather surprised-looking sheep on its cover, and it’s fair to say that her birth rocked the world.
The story of the technique that led to the creation of Dolly started more than 40 years earlier, back in the 1950s, when American biologists Robert Briggs and Thomas King started experimenting with frog cloning.
They were swapping out the nucleus (the biological bag containing DNA) from frog eggs and replacing it with a nucleus taken from a very early frog embryo. Yeah, there was a lot of frogspawn involved.
It’s important to point out that this wasn’t research aimed at cloning animals in a ‘Muhahaha! We’re playing God!!’ kind of way.
Instead, they wanted to test the idea first put forward by 19th century developmental biologist August Weissman, who suggested that cells in a developing embryo lost bits of their genetic information as they grew and specialised.
According to Weissman, the only complete, perfect set of DNA would be in a fertilised egg - any more mature cells would have lost the DNA they didn’t need and only kept the instructions for the cell type they had become.
This theory was backed up by Wilhelm Roux, who used a hot needle to obliterate one of the cells in a two-cell frog embryo, just after that very first cell division. The result was a very sad-looking half embryo, apparently proving Weissman right.
But another German embryologist, Hans Driesch, wasn’t convinced. He got busy shaking up sea urchin embryos in a test tube to the point where their cells fell apart from each other. Each one grew into a perfectly healthy new urchin, making these creatures the first true deliberately cloned animals in history, if you want to be pedantic about it.
These results showed that each embryonic cell still had all the correct instructions for life, suggesting that Roux had somehow damaged the remaining embryo cell with his hot needle experiments, preventing it from growing into a healthy frog.
Support for Driesch came from yet another German embryologist - Hans Spemann. Honestly, you’d think they were cloning them over there or something…
Spemann used a loop of hair from his own baby’s head to carefully separate the cells of a salamander embryo. Up until a certain time in development, which Spemann referred to as ‘determination’, any of the separated cells had the capacity to grow into new animals. But after that point, no dice.
Spemann’s salamander experiments were published in 1902, suggesting that it should be possible to clone animals from early embryo cells, but not from adult ones. That’s the law of nature, and that’s how it is. Case closed.
Briggs and King’s experiments in the 1950s took these German experiments to the next level, putting DNA from fertilised early embryo cells into unfertilised eggs, to see if it could direct normal development.
This was actually an idea first put forward by Hans Spemann in the 30s, but it took a while for embryologists to develop the tools and techniques that were needed for such a fiddly task.
By 1952, they had done it - successfully creating the first animal cloned by somatic cell nuclear transfer, as the technique was known.
Next came John Gurdon and his colleagues in Cambridge, who wanted to use the same technique to test the limits of Spemann’s point of no return and find out if it was possible to create cloned frogs using DNA taken from cells in the gut of older tadpoles, which were fully committed to their fate.
Amazingly they managed to get some live embryos out of all their experiments, some of which grew into healthy adults, netting Gurdon a share of the 2012 Nobel Prize in physiology or medicine for his work, together with Japanese stem cell biologist Shinya Yamanaka.
But while this proved that at least fully committed tadpole cells retained all their genetic instructions, Gurdon never managed to make a living clone from an adult frog cell.
Still he’d proved that amphibians could be cloned, so could the same technique be used to clone mammals?
There was an exciting false alarm in 1979, when University of Geneva Professor Karl Illmensee announced that he had managed to successfully clone three mice from early embryonic cells.
But when nobody else managed to replicate his results and a closer look at his lab revealed some strange goings-on, the university’s own enquiry judged that his results had been falsified - even though Illmensee himself still stands by his claim to have been the first person to clone a mammal.
And although people started having some success with cloning mammals after that – for example, successfully creating a cow using a cell taken from a very early embryo – it was widely thought that adult mammalian cells were somehow too far gone and set in their ways, biologically speaking.
It was impossible to turn the clock back. Until Dolly changed everything.
Dolly’s scientific dad, Ian Wilmut, entered the cloning game in the early 1990s, together with his colleague Keith Campbell at the Roslin Institute on the outskirts of Edinburgh.
As a leading centre for agricultural science, the Roslin researchers were busy using the latest genetic modification techniques on farmyard animals in order to boost yields and make them resistant to disease.
Having seen that cow could be cloned from early embryo cells – which can be grown in the lab and genetically manipulated – Wilmut wondered if this could be a way to make GM sheep.
Although Dolly gets all the glory, his first cloning successes were Megan and Morag. These were lambs cloned using stem cells that had been taken from a 9-day-old sheep embryo and grown in the lab.
The Roslin team’s success was due to a specific biological trick that they were using.
As cells grow and divide they go through a number of stages, known as the cell cycle. Along the way there are numerous processes and checks that are carried out, to make sure that the cell is ready to progress to the next stage.
Unlike previous cloning experiments, which had used donor cells from any stage of the cell cycle, Wilmut used cells that had been starved of specific nutrients, sending them into a kind of sleep.
Freed from the treadmill of the cell cycle, these dozy cells were much better DNA donors, ready to be reawakened in the egg and start development from scratch.
The birth of this pair and the growing proficiency of Wilmut’s team at the Roslin in manipulating these microscopic embryos led him to wonder whether cloning animals from adult cells really was biologically impossible, or it just hadn’t been done yet. So he took a leap from fetal cells to adult ones. But where to find them?
Next door to the Roslin Institute, a company called PPL Therapeutics were growing sheep breast cells in the lab as part of a project to try and genetically engineer sheep that would produce useful drugs in their milk.
The Roslin team procured some of the cells, put them to sleep, then began to pop the nucleus from each one into a waiting, empty sheep egg. If development started and the egg began to divide, they would transplant it into a ewe and see whether the pregnancy continued.
After nearly 300 attempts (because cloning is hard), they got just one lamb: 6LL3. This was the code number for the experiment that created her, and it was one of the livestock technicians who suggested the catchier name Dolly. And yes, she was named after Dolly Parton, because of the connection to boobs.
Then it all went a bit crazy. Dolly and her creators became scientific superstars overnight, although the work was reviled by some religious groups and animal rights activists. It’s an inevitable fact that if you want to clone an animal you’re going to have to break some eggs, and some people feel this is unacceptable.
It’s also a matter of some contention that neither Wilmut nor his closest colleague Keith Campbell won a Nobel prize for their work. Sadly, Campbell died suddenly just a week before John Gurdon’s award was announced in 2012.
It may seem like a bit of a silly question, I wanted to know what Dolly was actually like as an animal. It appears she shared some of the same outgoing personality traits as her namesake, as I discovered when I interviewed Ian Wilmut a couple of years ago.
“The best was to describe this is that I live down in the borders, in amongst sheep farmers. If they have a lamb whose mother has died, or she's got too many lambs, they take it into the house. It becomes accustomed to people and that's exactly what happened to Dolly.
“There were so many people visiting and wanted to see her, to get her to be in photographs, that she became accustomed to them. In actual fact she came forward to meet people when no ordinary farm sheep would do that - it would automatically turn and run.”
Despite her celebrity status, Dolly’s personal relationships managed to escape tabloid scrutiny. She had three sets of lambs by anonymous fathers – a singleton (Bonnie), twins (Sally and Rosie), and finally triplets (Lucy, Darcy and Cotton)
Unlike the real Dolly Parton, who is still going strong into her 8th decade, Dolly the sheep died at the age of six. That’s young for a sheep, which normally live for ten years or more. While there was much discussion about whether this was due to the fact she was a clone, there was a more depressing explanation.
Dolly caught a common virus that causes lung cancer in sheep, and after a few months she was suffering from untreatable tumours that made it hard for her to breathe. With much sadness, the Roslin team realised that only humane thing to do was to euthanize her.
Yet Dolly still lives on – not only in scientific legend, but also in corporeal form, stuffed and mounted in the National Museum of Scotland in Edinburgh. And she also lives in a much more real sense.
Four further clones were made from the original mammary cell line that gave rise to Dolly, a few years after her death. According to a paper published in 2016, Daisy, Debbie, Dianna, and Denise were healthy, happy nine-year-old sheep, living out their days in a university paddock.
To celebrate her 20th birthday, the Roslin Institute in Edinburgh where she was cloned and born decided to throw a party in her honour. Or rather, they had a scientific symposium, because scientists know how to have fun.
I was lucky enough to be invited to host the public event that evening, leading the assembled crowd in a noisy chorus of Happy Birthday to Dolly - possibly the only time that hundreds of people have serenaded a dead sheep.
Alas, I never did get to ask Wilmut whether he had a favourite Dolly Parton song. But I suspect that it’s probably, ‘I Will Always Love Ewe.’
References and further reading:
Healthy ageing of cloned sheep. K. D. Sinclair, et al. Nature Communications 7:12359 (2016)
Newkirk, Nicole, ""Transplantation of Living Nuclei from Blastula Cells into Enucleated Frogs' Eggs" (1952), by Robert Briggs and Thomas J. King". Embryo Project Encyclopedia (2007-11-13). ISSN: 1940-5030 http://embryo.asu.edu/handle/10776/1722.
Briggs, R. and King, T. J. “Transplantation of Living Nuclei from Blastula Cells into Enucleated Frogs’ Eggs.” Proceedings of the National Academy of Sciences 38 (1952): 455–463.
The first cloned animals were cloned over a century ago. Esther Inglis-Arkell, Gizmodo
Nottingham “Dollies” show that cloned sheep age normally, Roslin Institute
“Fertile” Intestine Nuclei. J. B. GURDON & V. UEHLINGER Nature 210, 1240–1241 (1966)
The Lie that Could Not Be Cloned, David Weiner (2015), Princeton Innovation Magazine
