For grand-prix competitor Mark ?Watring, of Hidden Valley, California, the opportunity for horse cloning was too intriguing to pass up. He had achieved international success?including an individual gold medal at the 2003 Pan American Games?with the Holsteiner gelding Sapphire. But he regretted that the champion show jumper, now 18, would never have the chance to produce offspring with the same athletic potential.
"Every rider has that special horse?the one they've had success on and built a special bond with," Mark says. "Most of them stay in the family?they aren't sold?once they're done competing. So you've got that 28-year-old winner out in the field, and when you look at him you can't help but wonder ?what if we could do that again?'"
In the not-too-distant future, Mark will be able to begin answering that question. After much investigation and research, he and his partners, John and Debi Bohannon, decided to clone Sapphire in 2009. Last February, Saphir, a colt who is genetically identical, was born using horse cloning. "I'm ready to ride him already," Mark says. "I'm very excited about it" and what the future may hold.
An Emerging Technology
Saphir is one of approximately 75 cloned horses who have been produced since the first equine clone?a mule named Idaho Gem?was born in May 2003 in the United States. Champions in cutting and barrel racing, former Olympic competitors, polo ponies, top Quarter Horse race winners and a Professional Rodeo Cowboys ?Association Horse of the Year have all been cloned. They are testimony to what can be accomplished through the advances in modern science. Yet the ethics of cloning still raises debate. And even among those who support its use, several important questions await answers. For instance, is a cloned horse truly identical to the original, and will he be able to demonstrate the same athletic ability?
ViaGen, the Austin, Texas, firm responsible for cloning Sapphire, is aiming to furnish answers through its work. The privately held company was founded in January 2002 to provide commercial bovine, equine and porcine gene banking, cloning and genomics services. In 2003 it acquired the rights to the cloning technologies developed by the Roslin Institute, the ?research facility in Edinburg, Scotland, where Dolly the sheep was cloned in 1996.
ViaGen cloned its first horse in 2006. Today, it is responsible for approximately 55 of the cloned horses living in the world, according to Candace Dobson, ViaGen marketing associate. Among them is Gemini?the 2008 clone of grand prix show-jumping legend Gem Twist. Among Gem's many lifetime accomplishments, the Thoroughbred gelding earned two silver medals at the 1988 Olympic Games and was named World's Best Horse at the 1990 World Equestrian Games in Stockholm, Sweden. He was cloned by his lifelong trainer Frank Chapot, who now owns Gemini and plans to stand him as a stallion. "The biggest part of our business is the geldings that people would like to have back as breeding stallions," Candace says.The Process
The cloning process at ViaGen begins with a $1,500 procedure known as gene banking to gather and preserve the genetic ?information found in an animal's DNA. According to Candace, a gene-banking kit is sent to a client's veterinarian. He uses the biopsy punch that it contains to extract a tissue sample about the size of a person's pinky fingernail from the crest of the horse's neck. The sample is then returned to ViaGen's lab, where a culture will yield millions of cells. They will be subjected to extremely low temperatures during a process called cryopreservation.
"Even if you're not ready to clone, gene banking is a simple process," Candace explains. "Once it's done, it's done," and it can be considered an investment in the future. For a fee of $150 a year, the preserved genetic material can be stored for an extended period. According to ViaGen, cellular DNA preserved in liquid nitrogen has been regrown after decades and could hypothetically be preserved for centuries.
"From that single sample, you could potentially clone indefinitely," Candace says. "You shouldn't ever have to recollect a horse." She adds that once a horse dies, it is usually too late to retrieve a viable sample. Tissue cannot be taken from a horse who has been euthanized.
When a client makes the decision to go forward with cloning, ViaGen takes an unfertilized egg (oocyte) from a donor mare and strips out the DNA. "You've essentially got a blank canvas," Candace explains. "Then we take one of the horse's preserved cells, insert it into the egg and fuse them in a process that imitates fertilization?the sperm hitting the egg." After that, the clone embryo starts to divide like a naturally conceived embryo. For a brief period it grows in a culture, then it undergoes conventional embryo transfer. "We put it in a surrogate mare," Candace says. "She carries it for about 11 months and out pops the baby."
ViaGen charges $165,000 for its cloning service: 10 percent is due when a contract is initiated, 40 percent when the recipient mare is 120 days along and the remaining 50 percent when the foal is 60 days old and ready to go home.
Complications and Misconceptions
As a way to foster understanding of the technologically advanced process, Candace emphasizes the importance of differentiating cloning from genetic engineering. "We're not changing any of the genetics of the horse," she says. "We're making a genetic duplicate. The explanation that people usually seem to understand is that a clone is an identical twin born some years later."
She admits that, like any other reproductive technology, cloning has its risks. "We're still in the early stages," she says. "Our efficiency rate right now is about on par for what embryo transfer was at the same age. If you think about it, the first clone in the world?Dolly the sheep?was born relatively recently, in 1996. So we're only about 15 years into it. When we do lose an embryo, it's usually early on, and it's simply reabsorbed by the mare.
"There are a lot of misconceptions out there about clones," Candace continues. "There have been research studies published noting difficulties such as birth defects and large umbilicus. ViaGen has simply not seen those same issues in our equine production."
A prevalent perception, according to Candace, is that cloning creates an unfair advantage. She disagrees. "There's so much environment that goes into a successful career. With cloning, you're looking at genetics. It's just another tool in the box.
"People with the most money already breed to the best horses." Candace continues. And there's no benefit in producing too much of a good thing. "Take Quarter Horse people, for example," she says. "They don't want to see a cutting class of 15 Smart Little Lenas. No one's ever going to do that. To make that many copies of a horse would only decrease its value."
Candace also responds to those who believe that cloning is not natural and compare it to playing God. "My answer is always that there's nothing about modern breeding that is natural," she says. "Selective breeding is not natural. You're interfering with the process just by selecting pedigrees to breed to."
ViaGen-cloned horses already have produced two generations of offspring through traditional reproductive means. According to Candace, there have been no negative repercussions. "We've had five or six horses who have a second foal crop on the ground, so they're out there producing normally and competing," she says. Still, "we don't have too many people ready to jump on the performance aspect of cloning because there is so much environment that goes into a horse's success." In fact, ViaGen recommends that its clients manage their expectations with regard to performance. "The only genetic guarantee that we can make is that the animal will have the exact same breeding value as the original. So please don't expect that Gem Twist's clone is going to go out and win an Olympic medal again."
An ?Uncertain Future
Katrin Hinrichs, DVM, PhD, is a professor at Texas A&M University's College of Veterinary Medicine and an equine cloning expert. In 2005, she led the team responsible for producing the first cloned horse in America, a colt named Paris Texas. She says there are several reasons why a clone may not be destined to have the same athletic ability as the original horse. "The first is simply environmental," she explains. "Perhaps the mother's milk isn't as good at birth. Or the youngster doesn't get the right nutrition or training. Things like that can happen to any foal."
A second potential hurdle is the newborn clone's overall health. According to Dr. Hinrichs, approximately 50 percent of the clones who have been produced by Texas A&M have been confronted with conditions early on that may have a lifelong effect. In her experience, "Many clones are born weak and have some problems at birth, such as contracted tendons or a large umbilicus that requires surgery," she says. "A foal that struggles for the first week or so of life may not turn out to be the individual he would have been," had he been born healthy. Dr. Hinrichs says that even a minor setback can make a significant difference in a foal's future. She adds that the mare's placenta?the organ that maintains and nourishes the fetus as it develops?plays an important role in the foal's health and is one of the tissues most affected by cloning. "It seems like a simple organ," she says. "But it is very complex, especially the way its development is controlled genetically."
Dr. Hinrichs points to one additional factor she believes is perhaps the most consequential in terms of its potential to compromise a clone's athletic ability: The current technology utilizes a skin cell to create the clone. "Although the nucleus of a skin cell has all the instructions to make everything in the body, it didn't use most of them while it was a skin cell. It wasn't using the genes that control the liver or the muscles or 99 percent of the body," she explains. "It was using only the DNA that is important when you're a skin cell."
As a result, the oocyte that has ?received the donor horse's DNA bears the responsibility for making a cascade of decisions to initiate and sustain embryonic development. "Essentially, the oocyte goes to the DNA and scratches its head and says, ?To make an embryo, I need this gene. But it has been turned off, and now I have to turn it on.' Or it discovers that another gene was turned on and now it needs to be turned off," Dr. Hinrichs ?explains. The oocyte goes through the entire set of DNA molecules?which carry some 50,000 genes?repeating the "need it, don't need it" process.
In many instances, the oocyte does a fine job of activating the necessary genes and deactivating others. But sometimes, the process is flawed. "If the oocyte does a really poor job, we won't get an embryo at all," Dr. Hinrichs says. "If it does it so-so, it won't make a pregnancy. If it does it pretty well, there likely is a pregnancy, but somewhere along the line, if there is a wrong gene that's available or shut off, this can cause the fetus to be lost. If, however, the oocyte does its work really well, the mare will go all the way to term and produce a foal."
In the case where a cloned foal ?ultimately is destined for breeding, what is the outlook for his or her offspring? According to Dr. Hinrichs, science indicates that, no matter how the clone itself is using its DNA, the clone's offspring should develop completely normally?no differently than any horse produced by traditional reproductive means. "The epigenetic markings?those that govern how the DNA is used?reset when an animal, a clone or not, makes eggs or sperm. These epigenetic markings do not carry on to the next generation," she says.
But a consideration that has not been well addressed, Dr. Hinrichs adds, is the fact that each oocyte contains mitochondria, the structures responsible for energy production that are found in every mammalian cell. Although mitochondrial DNA accounts for only 13 genes?a tiny fraction of the 50,000 genes contained in nuclear DNA?it cannot be stripped out of the oocyte. This means that some of the host egg's DNA?that is, the DNA contained in the mitochondria?is passed to the clone. "We don't know yet if it will have an ?effect on the clone itself. Remember, this is just a tiny amount of DNA, even the mitochondria itself gets most of its information from the nuclear DNA," Dr. Hinrichs says. "If the clone is a stallion used for breeding, there is no concern, because a stallion does not pass mitochondrial DNA to its offspring." But because the foal's mitochondria come from the egg, these tiny bits of DNA will pass to the next generation of a cloned mare.Registration Issues
Until more is known about horses who have been cloned, most equine breed groups are choosing not to register them. Among the first to address the issue was the American Quarter Horse Association, the world's largest equine breed registry and membership organization. Since 2004 its official handbook has included this rule: "American Quarter Horses produced by any cloning process are not eligible for registration." But the issue has been a source of ongoing discussion and debate.
At the association's 2008 convention, a change was proposed to the Stud Book and Registration Committee (SBRC). It involved allowing a live foal produced via cloning to be registered as long as its DNA matched that of a registered American Quarter Horse. At the time, the committee recommended postponing any decision pending further study. Later that year, representatives from ViaGen and educational research institutions met with the committee to discuss cloning and its ramifications.
The cloning rule change was again on the SBRC's agenda at the 2009 AQHA convention. There also was a cloning ?forum that included many industry experts. The SBRC recommended appointing a task force to seek information and input from knowledgeable sources regarding cloning and to conduct further study in four areas: parentage-verification issues, registration-process implications, general membership sentiment and implications with respect to genetic disease. Material gathered by the task force was presented to the SBRC at the 2010 AQHA convention last March. A member proposal to amend the rule regarding the registration of clones was discussed and then denied.
The Jockey Club, the breed registry for Thoroughbreds in the United States, Canada and Puerto Rico, is another organization that does not admit equine clones. Its rules for registration specify: "To be eligible for registration, a foal must be the result of a stallion's breeding with a broodmare (which is the physical mounting of a broodmare by a stallion with intromission of the penis and ejaculation of semen into the reproductive tract). As an aid to the breeding, a portion of the ejaculate produced by the stallion during such mating may immediately be placed in the uterus of the broodmare being bred. A natural gestation must take place in, and delivery must be from, the body of the same broodmare in which the foal was conceived. Without limiting the above, any foal resulting from or produced by the processes of artificial insemination, embryo transfer or transplant, cloning or any other form of genetic manipulation not herein specified, shall not be eligible for registration."
In contrast, the United States Equestrian Federation, the national governing body for equestrian sport, has no position on cloning nor does it place any restrictions on registering clones. However, many of its member affiliates, including The Foundation for the Pure Spanish Horse, the American Shetland Pony Club and the American Miniature Horse Registry, have developed their own position statements. According to The Foundation for the Pure Spanish Horse, "Until the registry is satisfied that it has gained a comfortable level of knowledge and assurances that specific technical, moral and legal aspects of cloning, gene splicing or other artificial attempts to enhance or manipulate the equine genome are resolved, the registry will not allow registration of any horses produced by such a manner."
A Look Ahead
In addition to serving as chairman of the USEF's Breeders Committee, Ruth Wilburn, DVM, of Olive Branch, Mississippi, breeds pure and part-bred Welsh ponies. She is also president of the Welsh Pony and Cob Society of America. She believes that associations based on performance or discipline will be more likely to adopt cloning than breed organizations. "If you have a wonderful event horse, you don't really care where it came from," she says. "But the breeds are going to look at cloning really hard because it can impact them so much.
"I don't think cloning has affected equestrian sport yet," she continues. "But it has the potential to affect us greatly if it becomes a common practice. There are a lot of unanswered questions. That's the big thing. For example, they need to figure out if these animals are going to be truly identical clones. After all, there is still a tiny bit of genetic material from the egg donor.
"There's also the matter of nature versus nurture," Dr. Wilburn continues. She ?believes that environment is a significant factor. "Say the cloned horse doesn't get the wonderful trainer who was such an integral part of the original horse's success. That will have a lot to do with it," she explains.
As Dr. Wilburn sees it, "We don't have enough animals on the ground yet to know everything we need to about cloning. We've learned a whole lot, but the more we think we know, the more we find out we don't know. Can they perform? What is that second and third generation of clones going to do? It's really an interesting subject, and people want to jump on the bandwagon. But with something so important that has the potential to affect a lot of breeds of horses?it's better to tread lightly," Dr. Wilburn says. "It's really hard to undo something like that if you make a bad decision. Then the mistakes are already in there and it's hard to get them out."