Genetics – a talk by Dr Jeff Sampson Reproduced with permission
In September 2000 Dr Jeff Sampson, the Kennel Club Canine Genetics Coordinator, gave a talk to the club on Genetics and Genetic testing. Genetic testing may be a future way forward to eliminate hereditary cataract in the breed. His talk is reproduced here with grateful thanks.
Genetics is very much a young science, as sciences go. The roots of genetics are to be found in the work of this monk, Gregor Mendel. Along with his duties as an abbot of his monastry, he also carried out experiments on breeding pea plants. He was particularly interested in the various forms that these plants could adopt, and just how the forms were inherited from generation to generation. He eventually published the results of his investigation in a scientific paper in 1856. Unfortunately the impact of this paper went largely unnoticed. This work was eventually ‘rediscovered’ in the very first years of the 20th. Century. Mendel’s results of his experimentation on pea breeding were set to form the foundations of a science called ‘genetics’, a term coined by William Bateson. During the remainder of the 20th. Century this fledgling science grew at a rate that far surpassed the rate of knowledge increase in the more traditional sciences. In 1953 James Watson and Francis Crick deduced the chemical structure of DNA, the chemical that makes up genes. From this point our understanding has increased at a rate that has certainly astonished even the more enthusiastic observers. The 21st Century begins with the prospect of shortly knowing each and every gene that is required to make a human, what the genes are, what they do, how they work and what happens when they don’t work properly. This information will be highly significant for the dog, because humans and dogs share very many of the same genes.
Let’s take a closer look at Mendel’s proposals. He made two very simple simple deductions: Firstly, he proposed that the characteristics of an individual, the way it looks and behaves, in fact everything about an individual, are inherited from its parents. He was the first to coin the term ‘gene’ and define it as a unit of inheritance and a determinant of a given characteristic. Secondly, he proposed that both male and female contribute to the characteristics of the offspring So, what Mendel was saying is that everything about an individual is inherited via the genes that it receives from both its mother and father.
Today we know a great deal about genes and how they work. The gene contains a genetic plan or blueprint that an individual requires to express a particular characteristic or trait. For example, the colour of your hair or eyes, how tall you are going to be, whether you can roll your tongue, in fact everything about an individual. Genes are made of s chemical called DNA and the plan is actually stored in the chemical structure of the DNA molecule that makes up a gene. Cells use the genetic plan embedded in a gene to make molecules called ‘proteins’ and it is the action of these proteins, working either individually or in groups, that actually determine the characteristics. The colour of your hair is determined by a group of proteins that manufacture the pigments that are responsible for hair colour. So it is the proteins, called enzymes, that make the pigments and determine hair colour, but an individual could not make the proteins necessary without first having the plans for their manufacture that are stored in the genes for these proteins. Current estimates suggest that we need somewhere in the region of 75,000 different genes (the word ‘genome’ is used to describe this collection of different genes) to produce a perfectly healthy dog. A similar estimate has been suggested for the genes required for a human. In fact, very many, possibly the vast majority, of the genes needed to specify a human are also required to specify a dog. Owners not only look like their dogs, they are also genetically very very similar to their dogs. One of the important features is that the dog possesses two copies of each and every gene in its genome.
Every cell in the dog’s body ( and there are something like one million million cells in total) contains two copies of each and every gene in its genome. The only exception to this are red blood cell, which actually possess no genes at all, and the reproductive cells, eggs and sperm, which only have one copy of each and every gene (see later). The genes are stored in a special area within the cell called the nucleus where they are assembled into structures called chromosomes. The dog has 38 different chromosomes (numbered 1 – 38) and these are shown above. Each chromosome carries genes along its length. The larger chromosomes , like chromosome 1 will have anything between 4,000 and 5,000 different genes arranged side-by-side along its length. Smaller chromosomes, like 38, carry fewer genes, say between 1,000 and 2,000. In fact, running along the length of each and every chromosome is a single molecule of DNA and different regions of this molecule actually represent the different genes. Remember, there are two copies of each and every gene and this means that there must be two copies of each chromosome. This picture quite clearly shows the different chromosome pairs; a pair of chromosome 1, 2 3, all the way down to a pair of chromosome 38. The dog also has a special pair of chromosomes that determine the sex of an individual. If a dog has two copies of the X chromosome, as is portrayed in this picture, it will be a female.
This picture shows the chromosomes taken from a male dog. It has two copies of each of the chromosomes 1 -38, just like the female seen before. However, its sex chromosomes are different. Instead of possessing two copies of the X chromosome it has one X chromosome and one Y chromosome. This is what makes it male. So, sex determination in the dog is just as it is in humans. Females have two copies of the X chromosome and males have one copy of X and one copy of Y. In order to distinguish these sex chromosomes from the rest we refer to them differently. Collectively, we call chromosomes 1-38 the autosomes; the X and Y chromosomes are called the the sex chromosomes. Let’s recap at this stage. Each and every cells in the female has a pair of each of the autosomes (chromosomes 1 – 38) and these contain the two complete copies of the 75,000 genes needed to make a dog. Every cell in the male has exactly the same complement of autosomes ( chromosomes 1-38) but now has an X and a Y sex chromosome. This has effectively covered the first of Mendel’s propositions; the gene as a unit of inheritance and determinant of a given characteristic or trait. What about his second proposal, that both male and female contribute to the characteristics of the offspring………?
The process of fertilisation results in the passage of parental characteristics to offspring. Bitches produce eggs and into each egg the bitch deposits one complete set of autosomes, I.e. one copy of each of chromosomes 1 – 38. Remember, each and every gene required for the dog will be present somewhere on one or other of these chromosomes. The egg therefore possesses a complete compliment of maternal genes. In addition, the egg contains just one copy of the X chromosome. Dogs, males, of course produce sperm. Each sperm head contains one of each of the males autosomes (chromosomes 1 – 38) and thus a complete set of paternal genes. However, each sperm head must also contain a sex chromosome. This means that there are in fact two different classes of sperm that can be produced. One will have the 38 autosomes and the paternal X chromosome, the other class will have a complete set 0of autosomes and the paternal Y chromosome. There will be approximately equal numbers of sperm in each class. At fertilisation the egg and sperm fuse and in this process the genes in the sperm head are injected into the egg which now becomes a fertilised egg. The fertilised egg now has two complete sets of autosomes and thus two complete sets of genes, one maternal and the other paternal. However, in terms of the sex chromosomes, there will be two types of fertilised egg. The egg fertilised by the sperm carrying an X chromosome will now have two X chromosomes and will develop into a female. The egg fertilised by the sperm carrying the Y chromosome will have one X and one Y and will develop into a a male.
This process of fertilisation explains Mendel’s second proposition and demonstrates how genes are passed from parents to their offspring. The fertilised egg contains two complete sets of chromosomes: a maternal set with one copy of each and every maternal gene and a paternal set with each and every paternal gene. This fertilised egg then ultimately produces the fully formed puppy. This requires a colossal amount of cell division. The fertilised egg is just one single cell, but it ultimately has to give rise to the million, million cells that are present in the puppy. The fertilsed egg initially grows and divides into two daughter cells. Each of these then grow and divide to give two of their own daughter cells, i.e. four cells in total. Each of these four divide to give eight cells in total, then 16, 32 64 and so on until the required cell number is achieved. Immediately before each and every one of these cell divisions every gene has to be copied so that the two daughter cells produced will still have two copies of every gene ( the maternal and paternal set) and two sex chromosomes. This represents a phenomenal amount of gene replication during the development of the puppy from a fertilised egg. Very early on in this process of cell division the cells produced appear to be identical and collected together into a ball of cells. However, soon, new cells produced by cell division take on different appearances and functions, a process known as cell differentiation. Cell differentiation is driven by the genes. Liver cells arise because they express a subset of genes that are required for liver cell to form; kidney cells express kidney genes, muscle cells muscle genes, and so on. Accompanying this process of cells differentiation and complicated cell movements so that slowly but surely the initial ball of cells grows and begins to take on the physical appearance of a puppy.
Sometimes the chemical structure of DNA within a particular gene becomes altered. Since the DNA structure stores the plan embedded in a gene, this alteration of a gene’s chemical structure will alter the plan. We call the alteration of the structure of DNA within a gene a MUTATION. Mutations essentially occur randomly in the DNA and so gene mutations are a random process and can in fact occur in any of the 75,000 different genes. Mutations are not common events so still the vast majority of genes in an individual will be unaltered.
We don’t know all of the causes of gene mutations, but we know of some: RADIATION. Most forms of radiation are known to cause structural damage to DNA and can lead to a gene mutation. For example, ultraviolet light in sunshine is hypothesised to cause gene mutations in some skin cells that can ultimately cause skin cancer. Other forms of radiation known to cause DNA damage include nuclear radiation. It is known, for example, that Russians who remained in the vicinity of Chernobyl following the nuclear accident there have 1000 times more mutations in their DNA than a control set of Russians that were nowhere near Chernobyl. CHEMICALS. Certain types of chemicals have also been shown to alter the structure of DNA and lead to gene mutations. Although both radiation and chemicals can be responsible for gene mutations, probably the vast majority of mutations come about as a natural consequence of the way that life has evolved. Two slides ago we discussed the colossal amount of gene replication that is required during the development of a fertilised egg into a puppy. This copying of the DNA structure within a gene is not 100% efficient and mistakes do occur. Thankfully the vast majority of copying errors are detected and corrected, but even this proof reading is not 100% efficient and, as a consequence, some copying errors go undetected and the DNA structure within a gene becomes mutated.
Mutations in genes do not necessarily spell disaster. In fact, the vast majority of gene mutations are neutral and don’t affect the plan embedded in the mutated gene. Sometimes the mutant gene affects the cell in which it first occurs. It could kill the cell or confer new, unwanted properties on the cell. Cancer is a good example of this. It is now generally believed that a cell has to accumulate mutations in a number of different genes before it becomes a cancer cell. It is known, for example, that 5 different genes need to be mutated before a colon cell becomes cancerous and gives rise to colon cancer. The mutations are in fact conferring the new property of immortality on the cell and it simply grows and grows in an essentially uncontrolled fashion. Such mutations are clearly critical for the cell; they are also critical for the animal in which the cell resides and can, ultimately cause the animal to die. But the mutation dies with the animal. The only mutations that are relevant for inherited disease are those that occur in genes where the altered plan causes a particular disease state and the mutant gene is put into either of the two types of reproductive cells, eggs or sperm. If a mutated gene that can cause a disease is put into an egg as part of bitches set of genes, or a sperm as part of the dog’s set of genes, then we will have the basis for an inherited disease.
It’s time to look at a specific example to attempt to draw together everything that I have been saying thus far about genes, inheritance, gene mutation and inherited disease. The example that I am going to use is that of sight, just one of the many characteristics of the dog. The biochemistry of vision in the dog is nearly identical to that in humans. The light that passes into the eye is focused onto the retina at the back of the eye. The cells that make up the retina are designed for just one purpose; to take this light information and convert it into a nerve impulse that passes down the optic nerve to the centre of the brain devoted to sight. There the nerve impulse is interpreted back to a picture by the neurons in the brain and we ‘see’ the image that the light entering the eye carried. We now know that inside each of the retinal cells is a group of proteins that are all required if sight is to be achieved. Each of these proteins will have a corresponding gene that is inherited at conception and used by the retinal cells to allow them to manufacture the protein. The number of different proteins required for sight is relatively large and could approach 50. There will be a corresponding number of vision genes that will need to function normally to allow the retinal cells to assemble a complete complement of retinal proteins. In this example I have chosen to reduce the complexity by assuming that only four different proteins are required ( proteins 1 -4), but remember this is a simplification to aid description.
The retinal cells contain a group of proteins ( 1- 4, for simplicity) that are required for vision. Normally these proteins are not active inside the retinal cell. You could imagine each protein to be a switch and all of them are switched off as a default situation. When the light signal hits the retinal cells it actually switches on protein 1 and activates it. This is shown by a change in shape here. Circular protein 1 is switched off and inactive. The energy in the light is used to alter the shape of protein 1, switching it on and giving it a new property. Once protein is switched on, it, in turn, switches on protein 2, i.e. activated protein 1 converts inactive protein 2 to active protein 2. Activated protein 2 then switches on protein 3 which then switches on protein 4. Once protein 4 is switched on a nerve impulse is generated that passes from the retina down the optic nerve to the brain. The generation of the nerve is represented here by switching on the light bulb. So, in summary, each retinal cell contains proteins required for sight and they all have to work in a precise sequence in order to generate a nerve impulse. The light activates the first protein in the chain, this then activates the second which then activates the third and so on until the final activated protein in this sequence generates a nerve impulse.
Now lets see what will happen if the gene for one of these vision proteins becomes mutated. Let’s assume that the gene for protein 3 in this pathway becomes mutated. I have represented this by changing the shape of protein 3 from a circle to a triangle. Let’s also assume the mutation actually alters the plan significantly so that retinal cells with the mutation actually make an altered protein 3 that cannot be activated. So, what happens now? The light will activate protein 1 which will activate protein 2. However, now protein 3 is altered so that it cannot be activated no matter how hard protein 2 tries. The sequence becomes blocked and because protein 3 cannot be activated, protein 4 remains in the switched off mode and a nerve impulse cannot be generated (the bulb remains switched off). The brain will therefore not know that light has passed into the eye. Sight will be lost.
Now let’s turn to looking at the different types of inherited disease. Broadly speaking, there are diseases that result from a single mutant gene and those that involve mutation of more then one different gene (polygenic) Single Gene Diseases Again there are two types of single gene disorders. To understand the difference you have to remember that there are two copies of each and every gene. When a mutation first occurs in a gene it only affects one of the two versions, the other gene copy remains normal. Once the mutation has occurred in a gene it will not be corrected and the mutant version will be passed on to future generations by breeding. One of the two types of disease mutation is called a recessive mutation and the other is termed a dominant mutation Recessive Mutations A disease caused by a recessive mutation will only ever be expressed if an individual inherits two copies of the mutant gene, one from its mum and the other from its dad. A dog that has one recessive mutant gene and a perfectly normal counterpart gene will not be clinically affected, but it will be a CARRIER. Dominant Mutations An animal need only inherit one copy of the dominant mutation to be affected. The dominant mutation is expressed even in the presence of a perfectly normal gene copy. So, to summarise, to be affected you need two copies of a recessive mutant gene, but only one copy of a dominant mutant gene. Polygenic Diseases Here more than one different type of gene is involved. The expression of the disease results only when an animal inherits mutations in more than one different gene. The classic example is that of hip dysplasia; we don’t know how many different genes need to be mutated for the disease to occur, but it is certainly more than one gene.
This slide represents the possible outcomes of a mating between two carriers of a disease caused by a single recessive mutation. The white circle represents the normal, non-mutated gene copy, the black circle the recessive mutant gene responsible for the disease. The bitch is a carrier and has one mutant gene and one normal gene copy (she is clinically clear). She will produce two types of egg, one carrying a full complement of genes including the normal gene and one carrying a full complement of genes but this time carrying the recessive mutant copy. The dog is also a carrier and produces two types of sperm, one carrying all the genes including the normal gene version and the other carrying all of the genes, but this time having the recessive mutant version of the gene. Remember, fertilisation is a random process so either class of egg can be fertilised by either class of sperm, so there are four different combinations that can occur. In this diagram I am attempting to use different colours to relate the outcome of a particular combination of egg and sperm at fertilisation. Consider the fertilisation of an egg by the sperm connected by the blue arrow. The puppy that develops from such a fertilisation is boxed in blue. The fertilisation depicted by a red arrow gives progeny that are boxed in in red, and so on. You can see from this that when you mate two carriers of a single recessive mutation you will expect to get 25% of the progeny that are both clinically and genetically clear (they have inherited a normal gene copy from both of their parents), 50% of the progeny that are carriers (these will still be clinically clear) and 25% of the progeny will be clinically affected because they have inherited a recessive mutant gene from both of their parents.
Now let’s do the same for a disease caused by a dominant mutation. Again, the white circle represents the normal gene copy and the black circle the dominant mutated version of the gene. You only need to have one copy of the dominant mutation to be clinically affected. This slide demonstrates the possible outcomes of mating an affected bitch (one normal gene copy and one dominant mutant gene copy) to a clear dog (two copies of the normal gene copy). The colour convention used here is exactly the same as described in the previous slide. Different types of fertilisation are related to the outcome of that fertilisation by virtue of using the same colour. You can see from this that if you mate the affected to a clear you would expect to get 50% of the progeny to be both clinically and genetically clear and 50% of the progeny will be affected because they have inherited their mother’s dominant mutant gene. A Note On These Expected Outcomes. The %s used in this slide and the previous slide are what we expect to happen in terms of how progeny will be distributed. However, I have stressed previously, fertilisation is completely random in terms of what type of sperm will fertilise what type of egg. This means that actual ratios of progeny in a litter may not be exactly what you expect. You expect from a carrier carrier cross to obtain 25% normals,50% carriers and 25% affecteds. These are simply probability estimates, in reality the distribution of pups in the litter might well be different. If you repeated the mating often enough and get a large number of puppies then the observed distribution would approach the expected distribution. However, in a single litter you could get an entire litter of clears, and entire litter of affecteds and anything else in between these two extremes. The %s quoted are simply a guideline.
This slide depicts the theoretical outcomes of matings between dogs that may or may not have a recessive mutant gene that causes a clinical disease. The symbols used here are new and therefore need some explanation: Squares represents males (dogs), circles represent females (bitches). A horizontal line connecting a dog and bitch represents a mating and the progeny that come from such a mating are connected by a vertical line. The symbols can be split to represent the genetic makeup. So, of the symbol is completely white the animal has two normal versions of the gene and is genetically and clinically clear. If one half is white and the other black, the animal has one recessive mutant copy and one normal copy of the gene. It is a carrier and will be clinically clear. A wholly black symbol represent an affected because it has two copies of the recessive mutant gene. Obviously, clear to clear is ideal and all offspring will be clear, both clinically and genetically. A clear mated to a carrier will give expected ratios in the progeny of 50% clears and 50% carriers. We have already seen the outcome of a carrier carrier mating. Mating an affected to a clear will give you an entire litter of carriers (this will always occur and doesn’t depend on probabilities). If you mate a carrier to an affected you would expect to get a litter in which 505 are carriers and 50% are affecteds. Finally, mating two affected can only give you an entire litter of affecteds.
This slide shows the expected outcomes from various matings for a disease caused by a dominant mutation. Remember, you only need one copy of the dominant mutant gene to be clinically affected. On the top we see the outcome of a clear to a clear, as expected all progeny will be clear. Moving to the second row, on the left, if an affected is mated to a clear you would expect half the litter to be clear and half to be affected. This is assuming that the affected only has one dominant mutant version of the gene. Occasionally, but a lot less common, an affected will actually have two copies of the dominant mutation. If such an affected is mated to a clear the outcome will be different (second row, right hand side). In this case all of the puppies will be affected. The bottom row depicts the outcome of mating an affected to an affected. On the extreme left we what happens if both of the affecteds have a single copy of the dominant mutation. You would expect to get 75% of the litter affected, but 25% both clinically and genetically clear. The other two examples show what happens if one or both affecteds actually have two copies of the dominant mutant gene. In both cases (bottom row middle and extreme right) all of the progeny will be affected.
Inherited Disease in the Dog
Now let’s look at the types of inherited disease that have been recognised in the dog.
There are now 370 recognised inherited diseases in the dog. This sounds like a large number, but contrast the same estimate in humans where there are now in excess of 4000 different inherited conditions. Of the 370 canine inherited diseases, 167 have yet to completely defined and we have no information on their precise mode of inheritance. Of the remaining 203 conditions where we know the mode of inheritance, 66% are the result of a single autosomal recessive mutation, 15% by an autosomal dominant mutation, 5% by an X-linked recessive mutation and 20% are polygenic. The only class of mutation that we haven’t yet covered are the X-linked mutations. These refer to mutations in genes that reside on the chromosome, one of the chromosomes that determines the sex of a dog. Diseases caused by mutations of genes on the X chromosome have a very characteristic mode of inheritance. The disease is normally passed down the female but is usually only expressed in male offspring. Consider a recessive mutation of a gene on the X chromosome. The gene causing haemophilia A is one such X gene. Females that have a haemophilia mutation on one of the X chromosomes are not normally affected because the condition is recessive and their other X chromosome will have a normal copy of the gene. So, they can either pass the X chromosome containing the disease gene to their son or their other X chromosome containing the normal gene copy. Sons receiving the maternal X carrying the disease gene will become affected because their other sex chromosome is Y, inherited from dad, and it will not have a normal copy of this mutated gene.
The problem with data like that provided in the previous slide is that it is largely gathered by veterinary specialists who see nearly all of the affected cases. Of course, unaffected dogs don’t go to the vet, so it is not really possible to say just how relevant a condition is to a particular breed. It is not sufficient simply to say that an inherited condition has been reported in a particular breed, we need to also know how frequently the condition occurs within a breed. Breeders are clearly ideally placed to provide some insight into the prevelance of a given condition within the breed. They are the ones that breed the puppies and they are the ones that will know when puppies are affected with an inherited condition and how frequently such puppies appear. It is therefore important that breed clubs and councils have health surveillance in place in order that they can gather information on disease prevelance. From a recent questionnaire designed to discover just how many breed clubs or councils in this country have some kind of health surveillance, we estimate that only around 20% of the clubs and councils in this country have established health surveillance. It is important for those clubs that haven’t done so already to establish either a health coordinator or a health committee.
This slide shows the kind of things that those involved with health surveillance in a breed can do and the sort of information that they can provide in order to help breeders construct better breeding programmes that might limit the spread of inherited disease. The information above is fairly self explanatory and so we don’t need to elaborate any further.
It is important that breeders screen their breeding stock before they are used for breeding so that they have a better idea of what genes they carry and what matings are likely to be compatible and avoid producing affected offspring. This is not an exact science because we cannot yet test for many of the genes that cause inherited disease. However, it is certainly possible to ascertain whether a dog is clinically affected with an inherited condition. Since many of these will result from recessive mutant genes, the identification of a clinically affected individual immediately identifies both of its parents as carriers of the condition (provided the disease is caused by a single recessive mutant gene). The parents of dogs that are clinically affected with a polygenic disease are also suspect, because both must contribute to the clinical status of their offspring, although not necessarily equally. Health screening breeding stock should become an accepted part of breeding and all breeders should be encouraged to check their breeding stock before they use them for breeding. The results of health screening and their incorporation into breeding programmes can make a significant contribution to limiting the spread of inherited disease to future generations of dogs.
There are three health screening programmes that have been developed and run jointly by the Kennel Club and the British Veterinary Association: The BVA/KC/ISDS Eye Scheme: Checks for inherited conditions that affect the eye. The BVA/KC Hip Scoring Scheme that checks for the quality of a dog’s hips in relation to Hip Dysplasia. The BVA/KC Elbow Scoring Scheme that checks for Elbow Dysplasia
Identifying dogs that are affected with an inherited disease and not breeding from them will certainly, over a period of years, significantly reduce the incidence of the disease within the breed. For conditions that result from a dominant mutation, not breeding from affecteds will essentially remove the disease gene from the breed’s gene pool. However, most inherited disease in the dog results from recessive mutations. Unfortunately just removing affecteds from the breeding programme will have significantly less effect on the overall breed incidence of the condition. This is because health screening does not identify carriers. Carriers are usually perfectly normal clinically. Removal of affecteds will reduce the carrier frequency, but not to the same extent as the affecteds will go down. These carriers act as a silent reservoir of the mutant disease gene. The only way to make significant in-roads into recessively inherited disease is to be able to identify carriers and take note of this information when constructing breeding programmes. What is needed is a really good way of identifying carriers. This could be done if we were able to identify the genes responsible for inherited disease. Once we can identify individual mutant genes, simple DNA tests of breeding stock will tell us whether they are genetically clear, a carrier or an affected.. This information will then be invaluable to the breeder who, over a number of generations, could remove the mutant gene from their line.
The most straightforward way of identifying genes responsible for inherited disease, although not always possible, is the CANDIDATE GENE APPROACH. Many inherited disease in dogs have equivalent diseases in man, mouse and other species. When scientists discover genes that cause disease in these species, this often acts as a clue as to the dog gene involved. Two examples should suffice to illustrate the power of this approach. PRA IN THE IRISH SETTER is clinically very similar to a human disease called Retinitis Pigmentosa (RP) and a blinding disease in mouse caused by a mutation known as rde. When it was discovered that a mutation in the same gene caused one form of RP in man and the rde condition in mouse, scientists checked to see if the same gene was involved in PRA in the Irish Setter, and it was. The gene responsible for PRA in the Irish Setter had been identified and a DNA test for this mutant gene quickly followed. CLAD IN THE IRISH SETTER is an inherited disease where affected dogs have a severely compromised immune system. CLAD stands for Canine Leucocyte Adhesion Deficiency. Humans (LAD) and cows (BLAD) suffer from a very similar inherited condition. The same gene mutation causes LAD and BLAD and this information allowed us to check whether the same gene was involved in CLAD in the Irish Setter. It was and so a DNA test for CLAD quickly followed. Irish Red and White Setters also suffer from CLAD and the DNA test developed for Irish Setters was also shown to work for the CLAD gene in this breed.
Identifying candidate genes in other species is not always going to be possible. We therefore need another way of identifying mutant, disease genes. The recently developed Canine Genome Map will be an invaluable resource to this end. The Genome Map has resulted in easily identifiable marker DNA sequences being placed along each and every chromosome in the dog. Each marker uniquely identifies just one region of a chromosome. So, how will this help to identify disease genes? Well, although we don’t know where a gene is located, we do know that it will be on one or other of the canine chromosomes and will have one of the markers that make up the genome map on either side of it. If we could identify the DNA markers on either side, we will have located the gene to a relatively small area of just one of the chromosomes. It then becomes a much simpler task to look at the genes known to be present in this newly identified chromosome region and see if any are likely to be candidate genes for the disease. Identifying the markers that flank a gene is relatively straightforward and relies on the fact the regions of DNA close to each other on the same chromosome tend to be inherited together. By analysing DNA from individuals from a pedigree where one or more of the offspring are affected we can identify the markers that lie either side of the disease gene.
The markers that lie on either side of a disease gene are said to be LINKED to the disease gene. As we saw on the previous slide, identifying markers linked to the disease gene locates the gene to one small region of one of the chromosomes. However, the identification of linked markers allows the development of a DNA test for the disease gene. Essentially, dogs that have the linked marker are very likely to have the disease gene as well. So, a linked marker test can be developed once the linked markers are known. Such tests can be of great value to the breeder, but they are not 100% accurate, a fact that also needs to be taken into account. However, if used carefully, breeders can use linked marker tests to begin to construct better breeding programmes. Eventually linked markers will develop into Gene based tests, because the linked markers will greatly accelerate the discovery of the disease gene. Candidate gene approaches immediately identify the disease gene and lead to the immediate development of gene based tests. These tests are the Rolls Royce of DNA testing because they have 100% accuracy.
The great value of having DNA tests available for disease genes is that dogs can be prescreened before they are used for mating. Breeders will know that there dog is clear (two perfectly normal gene copies), a carrier (one normal gene and one mutant gene) or is affected (two copies of the mutant gene) before it is used for mating. This information will greatly improve breeding decisions. For example, breeders will be able to avoid mating carriers to carriers and the possibility of producing affected offspring.
Once a DNA test is available for the disease gene is available, breeders will be able use their carriers for breeding. This is vitally important. In the past carriers have been identified by other means. But the advice has always been not to breed from carriers. Whilst this has to be the best genetical advice, it might not be the best for the breed as a whole. Not breeding from carriers will certainly halt the further spread of the disease gene, but it will also deny the breed the possibility of inheriting the positive features and genes that a carrier might provide. By not breeding from any proven carriers could well be throwing the baby out with the bath water. An available DNA test for a disease gene makes the mating of carriers possible. So, that really good specimen that just happens to be a carrier can now be bred from and the breed as a whole can benefit from the positive things the dog has to offer. DNA testing allows carriers to be bred because it allows the owners of the carrier to choose a genetically normal partner. If a carrier is bred to a clear, approximately half the litter will be carriers and the other half will be clear. Furthermore, the DNA test can be used to screen the puppies to identify the carriers and the clears in the litter, allowing clear progeny to be bred on from.
In the absence of an available DNA test, breeding from known affected should be discouraged. However, once a DNA test is available, breeding from affected should be possible under appropriate circumstances. Again, the DNA test will allow the owner of the affected dog to choose a genetically clear mate. All of the litter will be carriers, but one of the bitch puppies could eventually be mated to another clear dog and this second generations will be approximately 50% clear and 50% carrier. Again DNA testing the progeny in this second generation will identify clear puppies that can eventually be used to extend the line to future generations.
The Kennel Club recognises the potential role that it can play to encourage the use of future DNA testing and the subsequent reduction in the disease burden imposed by inherited disease. It has accepted that the results of DNA testing can be incorporated into the registration process. Once a gene based test becomes available, the Kennel Club, in collaboration with the breed clubs and councils concerned, will draw up a Control Scheme for the condition. The control scheme for CLAD in the Irish Setter is given above as an example. Essentially it states that following the introduction of a new gene based test for a mutant gene, the breed should very quickly move to a situation where all breeding stock are DNA tested. With such information, breeders will then be allowed to breed their carriers provided that a carrier is only mated to a clear dog. If such a mating takes place, all progeny should be DNA tested to identify the carrier puppies and the clear puppies. Breeders will have a window of time, 5 years was the window decided by the Irish Setter breeders, during which carriers can be mated to clear dogs. However, after this window of time has elapsed, dogs will only be registered if they can be shown to be clear of the condition, either by direct DNA testing, or by virtue of the fact that both their parents were tested clear, I.e. the dog is hereditarily clear. After this window of opportunity has closed, no carriers will be registered by the Kennel Club. This will form the model for all future gene based tests, provided that the breed clubs and councils agree. The only major difference will be the window of opportunity during which carriers can be mated to clears. The Irish Setter breeders decided that 5 years would be sufficient. Other breed clubs might decide that this window should be either lengthened or shortened as appropriate. This type of breed control system will not be invoked by the Kennel Club if the DNA test developed is of the linkage type. This less accurate test is not considered robust enough to base decision of registration on.
There are only DNA tests available for a relative handful of inherited disease. Much more research is required to develop new breed-specific DNA tests for inherited disease. This inevitably, means that more research will be needed, research that will somehow need to be funded. That is why the Kennel Club has recently established the Kennel Club Health Foundation Fund. Formally part of the Kennel Club Charitable Trust, the Health Foundation Fund will provide ring-fenced funding to allow breed clubs and councils to develop further DNA tests that will benefit their breed. The idea is that clubs and councils will raise part of the funding toward the cost of a research programme and the Health Foundation Fund will provide top-up funding to allow the research programme to go ahead. The precise level of support raised by the club will not be an issue. In this way it is hoped that the numerically small clubs will have as much access to top-up funding as the numerically larger clubs with their increased ability to raise funds of their own.