Post on 12-Jan-2017
Breed Improvement Through DNA Testing By Nathan Dewsbury, MSc
Breed stewards and enthusiasts all care about the health and greater well being of their animals. All animal lovers want to provide the best for their pets in the way of the best food, the best health care, and the best toys available. Breed stewards seek to go beyond the external applications and instead look internally at the genetic aspect of pampering their animals in the form of genetic improvement. It’s this fundamental desire of providing the best for another life form other than your own that leads breed stewards to take on the task of breeding. A steward, who is already knowledgeable in their breed of choice, is always aware of the genetic health conditions of their breed and serves as the most commonly sought aspect of improvement. This increased search for greater canine health is driving the development for better tools, in the form of DNA testing that is going to give breed stewards a “leg -‐up” in breed development. So What is Merle and Why is it a Problem? In the domestic dog merle coloration is found in a number of breeds (Australian Shepherd, Cardigan Welsh Corgi, Shetland Sheepdog, Collie, Great Dane, Catahoula Leopard Dog) recognized by major canine registries. It is in these breeds, where the merle phenotype (coloration) is common place, that stewards are faced with a phenotype that is linked to major physical defects most notable in eye sight and hearing impairment. Both of which defects drastically affect the daily life and the over all well being of the affect animal(s) from birth.
Merle coloration is presented as patches of diluted pigment interspersed with normal melanin coloration of that breed, which usually shows up as spots (Figure 1). This merle phenotype is the result of a mutation that has resulted in additional genetic information being inserted into the normal (wild type) gene called SILV, which is found on chromosome 10. This additional genetic code is the result of a class of Retrotransposon, referred to as a jumping gene, that can cut and past it’s self any where in the canine genome, thus altering the function and expression if inserted in a gene. The SILV gene itself has a central role in pigmentation with significant expression of the gene in the skin and the eye. This functional role is especially crucial as lack of pigmentation in the hair inside of the ear canal results in deafness. Most individuals know the association of pigmentation and its link to genetic abnormalities on a general level. Skin color in humans is a good example. We know that the lighter your skin complexion the greater your possibilities to develop certain skin cancers if unprotected under prolonged exposures in the sun. Merle pigmentation is determined during embryo development, not a result of an onset condition that could possibly be treated or reversed. The merle mutation is responsible for developmental abnormalities that affects cardiac, ophthalmic, auditory, and reproductive systems in these domestic canines. In the case of the merle mutation, the developmental issues exhibited by affected canines share similar abnormalities observed in the human auditory-‐pigmentation disorder Waardenburg syndrome (WS). Both merle and WS can
display a range of physical changes as a result of the genetic changes. Alterations include the appearance of blue eye(s), eye shape deformities, blindness, levels of deafness, and cardiac valve problems.
Figure 1. Demonstrates the three possible phenotypes associated with the semi-‐dominant inheritance of the merle phenotype. The dog on the left contains a double merle, in the middle is a normal (wild type) dog, and the dog on the right inherited a single merle gene. Image taken from Genomics of Disease book
chapter (Stadler Genomic Symposia Series) by J.P. Gustafson, March. 2008
How is the Merle Mutation Passed On?
Knowing the deleterious effects associated with the merle genotype one might ask how is it that the merle pattern, hence the merle mutation, is still present in certain breeds? Breeding practices alone, no matter how stringent the selection criteria, could not selectively breed out the mutation with out genetic testing. This is the primary reason for the continual occurrence of the merle mutation after the divergence of canine breeds and still today. In addition to the possible expressed phenotypes (Figure 1) the situation arises when a dog can be genotypic merle yet does not present a traditional merle pattern phenotype. These dogs are coined a cryptic merle. These dogs appear to have solid coat colors or multiple solid colors instead of the obvious merle-‐spotted appearance. It was assumed that these dogs were solid or non-‐merle dogs and thus would be incorporated into the breeding population only to perpetuate the merle mutation. The only way to fully know the genotype of a dog is through DNA testing.
The merle mutational trait is inherited in an autosomal semi-‐dominant fashion as demonstrated by the hypothetical breeding illustrated in Figure 2 below. Using a Punett square model, we can see in Figure 2, that the presence of a merle genotype parent alone is not enough to produce all merle offspring. The semi-‐dominance is expressed in terms of the presence of a single merle allele that when inherited will be expressed dominantly in the phenotype of that offspring, but in the absence of the mutant allele the wild type phenotype will be expressed dominantly (Figure 2, Breeding B & D) even if one or both parents are carriers. Inheritance of a single merle locus (heterozygous) usually is not associated with any health effects
for that animal. In all merle breeds that are genotypic double merles (homozygous) for the merle locus will result in subleathal effects. This genotype is responsible for the multiple abnormalities associated with the incorporated merle mutation. An exception to the resultant double merle effects is found in the Catahoula Leopard Dog. The merle mutation has shown to behave differently with this breed, which shows less susceptibility to exhibit the negative effects of merling when compared to other breeds.
Figure 2. Demonstrates the breeding possibilities to illustrate the semi-‐dominant inheritance pattern of the merle gene found in the domestic canine.
How Can DNA Testing Help?
Once only imaged by breeders as an act of science fiction, DNA diagnostic tests are becoming increasingly available through DNA testing services and may soon become the norm in future breeding programs or even in pet selection.
The use of DNA diagnostic tests have been implemented for a number years and are primarily conducted or initiated by veterinarians for the diagnosis of ailments and inflictions such as canine epilepsy or Parvovirus. Older diagnostic methods entailed multiple complex steps that lengthened reporting of results yielding semi-‐specific results. Most diagnostic tests are implemented in the detection of an on-‐set health aliment. New innovations in research have enabled the development of more streamline testing protocols yielding specific results with fewer steps. An additional aspect brought about by innovation is the use of DNA. The ability of a diagnostic approach to screen DNA is now preventing abnormal health development via detected genetic conditions in parental animals that would have caused or led to health defects. The advantages of DNA diagnostics are
demonstrated by the availability of diagnostic tests to determine parentage, presence of specific disease markers, prediction of coat coloration and length are to name a few offered by companies outside of the veterinary care system.
Now stewards faced with the merle phenotype have yet another tool to
further breed health progression with the development of the Merle Test, a DNA diagnostic test, for the detection of the merle mutation. Prior to the development of merle testing, such enacted breeding practices to diminish the merle occurrence and detrimental affects entailed: no merle-‐to-‐merle breeding, breeding merle to only solid colored dogs, avoidance of predominant white dogs in breeding programs, and scatter breeding to solid colored dogs every two or three generations for certain breeds. These practices have served breeders well, but they take a large investment in time, breeding, and experience to prove the level of merle expression. This leaves error of selection and opportunity for vast improvement in way of breed improvement.
Dr. Leigh Anne Clark, Texas A&M canine geneticist developed the licensed
Merle Test during her research in determining merle patterning in the domestic dog. This straightforward DNA diagnostic test, provided by GenMark’s Merle Test, determines the presence of the merle gene and it’s level of genetic dominance. A dog owner wishing to know the genotype of their merle dog or those associated with merle dogs have to simply take a mouth swab of the animal(s) of interest and send the swab back to GenMark for testing and analysis. Using a polymerase chain reaction (PCR) method allows GenMark to amplify the region of chromosome 10 that encodes for the SILV gene, which will either contain a mutational insert or not. The results gained from this testing informs the owner(s) to the level of merle expression in their animals as carriers, non-‐carriers or genetically dominant (double merle) for the merle locus.
Armed with conclusive evidence allows for more informative breeding
practices and improved mate selections in an effort to remove the merle mutation from breeding populations of domestic canines. The decreasing presence of the merle mutation will lead to an increase in breed health. Eventual removal through DNA testing and selective breeding practices will result in breed improvement on the basis of removing an un-‐advantageous genetic source that, when present, only introduces defects in important functional systems of life. Advances in DNA diagnostics, through further understanding of the canine genome, in the form of genetic inheritance testing for animals is proving to be a real world tool available to people. DNA diagnostics has the ability to translate scientific research into functional applications to aid breed stewards in their efforts toward canine health improvement.