by Kristin Berkery

go to part IV | part I | part II

The previous I and II installments of Makin’ Babies examined how artificial insemination, transported semen, and embryo transfer can influence breed trends.

Pieraz, the first horse to be cloned.

There’s one breeding technology that isn’t approved by registries in the U.S. but has been used in Quarter Horses, Arabians, and Warmblood breeds — cloning.

One of the first horses to be cloned was an Arabian, the accomplished endurance racing gelding, Pieraz. Because Pieraz can’t be used for breeding, he was cloned so his genes could be passed on to the next generation. Pieraz’s clone, called “Pieraz Cryozootech Stallion,” stands at stud in France. The stallion clone was registered by the Anglo European Studbook, an organization that registers performance horses for competition.

LD Pistal, the double National Champion Senior Stallion who was cloned. Photo by Kelly Campbell

In 2007 a colt who was a clone of the 2006 and 2008 U.S. National Champion Senior Arabian Stallion, LD Pistal, was born.

Horse clones are produced through somatic cell nuclear transfer (SCNT), a process that involves taking a tissue sample from a horse, extracting the DNA from it, and inserting the DNA into an egg cell that has had the nucleus removed. The egg is stimulated to start dividing into a blastocyst, or early embryo, and then implanted in a surrogate mare who carries it to term.

The donor eggs used in the cloning process are usually from ovaries harvested from slaughtered mares. The horses are not slaughtered specifically for their ovaries; the harvested ovaries are a by-product of processing the animals. Since horse slaughter has been banned in the U.S., cloning facilities must get their donor eggs from other countries. One cloning company, ViaGen, built a facility close to a Canadian slaughterhouse to lower the cost of obtaining donor eggs.

A diagram of SCNT cloning. Therapeutic cloning means the resulting somatic cell is used for regenerative medicine, i.e. growing tissue for burn victims. Diagram by Dr. Jürgen Groth. Click the image for a larger view.

The reason LD Pistal was cloned was because he is subfertile and his clone would replace him in the breeding barn. There is one major roadblock: The Arabian Horse Association doesn’t allow clones to be registered or used for breeding registered horses. After the foal was born, its owner, Jeff Sloan, requested to meet with the registration committee to discuss whether cloning might be permitted by the AHA in the near future. The meeting was not held however, as LD Pistal and his clone were sold as a package to a new owner, Felix Cantu. Sloan chose not to pursue the issue any further, leaving it to Cantu to advocate for clone registrations.

It’s popular to say that a clone is “genetically identical” to the “parent” that donated its DNA to the “child,” but this isn’t actually true in the case of SCNT. According to Wikipedia:

In SCNT, not all of the donor cell’s genetic information is transferred, as the donor cell’s mitochondria that contain their own mitochondrial DNA are left behind. The resulting hybrid cells retain those mitochondrial structures which originally belonged to the egg. As a consequence, clones such as Dolly [the sheep] that are born from SCNT are not perfect copies of the donor of the nucleus.

Mitochondrial DNA is only passed from mother to child, and since there’s not actually a “mother” contributing her DNA to the foal, that missing genetic information is being pulled from the egg that was used in the cloning process. According to studies done on sheep cloned through SCNT, no mitochrondrial DNA from the donor (the animal that was cloned) was present in the resulting cloned offspring. This means the offspring obtained through SCNT cannot be genetically identical to the animal that was cloned.

Dolly the sheep.

The first successfully cloned animal, Dolly the sheep, died at the age of seven after a respiratory infection. (Sheep life expectancy is about 10 to 12 years.) In a report published in Human Molecular Genetics, studies have shown human mitochondrial DNA that has mutated tends to give rise to respiratory defects and age-related disorders, suggesting that SCNT clones will not live as long or be as healthy as their donor “parent.” The subject is not completely understood by scientists, but these studies serve as a warning that when mitochrondrial DNA becomes mutated, the resulting cloned animal may have health issues.

Pieraz clone horse

Pieraz’s clone. Photo from The Horse Magazine.

Not only are SCNT clones not genetically identical to their donor “parent,” they’re also not physically identical. Just like identical twins, there are small traits like white markings that appear different on a clone, which is a result of various cells migrating during gestation.

Should registries allow cloned animals to be registered for breeding purposes? There’s one immediate argument against it: Clones obtained through SCNT are not genetically identical to the original animal. With stallion clones used for breeding, they don’t pass on their mitochondrial DNA because the mares they’re bred to contribute their own mitochondrial DNA, essentially cancelling out the “foreign” mitochondrial DNA that comes from the donor egg during the cloning process.

Compare the markings on show jumper ET (left) with his clone (right).

But mare clones will not “breed true” — since their mitochondrial DNA comes from the donor egg during cloning, mare clones will pass on “foreign” mitochondrial DNA to their foals. The only way around this issue is to use a donor egg in the cloning process that comes from the same mare being cloned, but that requires the donor mare to be alive and still producing eggs. This work-around would also add to the cost of the procedure since it’s more complicated to harvest viable donor eggs from a living horse.

The point of a registering authority is to keep accurate records of all horses’ pedigrees, but how can parentage be determined of a horse that wasn’t the product of conception and carries the genetic material of an unknown donor? A great emphasis is placed on “purity” in Arabians, and the issue of SCNT introduces more purity questions.

read more in part IV | part I | part II

(You may leave your comments on part IV.)