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  • Author or Editor: Kunal Ray x
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To develop a molecular diagnostic test to ascertain genotype of the mucopolysaccharidosis type VII (MPS VII) locus.

Sample Population

Blood samples from 45 mixed-breed (German Shepherd Dog X Beagle) dogs that were phenotypically normal or affected with MPS VII.


The canine ß-glucuronidase gene (exon 3) was amplified by polymerase chain reaction (PCR), using 2 pairs of primers to determine the genotype of the MPS VII locus by 2 independent methods. For the first method, PCR products were used for heteroduplex analysis, using conformation-sensitive gel electrophoresis. In the second method, an allele-specific restriction site was created by use of a mismatch primer in PCR. The amplified DNA fragment was digested with a restriction enzyme (Eag I) to enable identification of the wild-type and mutant alleles.


Conformation-sensitive gel electrophoresis resulted in a single DNA band representing homoduplex when the sample contained a wild-type or MPS VII allele, but 2 bands representing hetero- and homoduplexes when both alleles were in the sample. Restriction digestion of the DNA fragment obtained by use of a mismatch primer was cleaved only when the template was a wild-type allele. Thus, samples from phenotypically normal carrier dogs that contained wild-type and MPS VII alleles were partially digested by the enzyme.


The diagnostic test used 2 strategies for independently ascertaining the wild-type or mutant MPS VII alleles in dogs. Thus, test results can distinguish phenotypically normal MPS Vll-carrier dogs from homozygous normal dogs. (Am J Vet Res 1998;59:1092-1095)

Free access
in American Journal of Veterinary Research


Objective—To define the relationship between clinical expression of a type-1 von Willebrand disease phenotype and genotype at 2 von Willebrand factor marker loci in Doberman Pinschers.

Animals—102 client-owned Doberman Pinschers.

Procedures—Dogs were recruited on the basis of plasma von Willebrand factor concentration, clinical history, and pedigree. Blood samples and response to a history questionnaire were obtained for each dog. Plasma von Willebrand factor concentration was measured by use of an ELISA, and genotyping was performed via polymerase chain reaction for 1 intragenic and 1 extragenic von Willebrand factor marker. Amplification product size was determined by use of polyacrylamide gel electrophoresis (intragenic marker) or automated sequence analysis (extragenic marker). Western blots were prepared from a subset of dogs with low plasma von Willebrand factor concentration to evaluate multimer distribution.

Results—Strong associations were detected between plasma von Willebrand factor concentration and von Willebrand factor marker genotype. Twentyfive dogs had substantial reduction in plasma von Willebrand factor concentration and multiple hemorrhagic events. All were homozygous for a 157-basepair intragenic marker allele and homozygous or compound heterozygous for 1 of 4 extragenic marker alleles. These marker genotypes were exclusively detected in dogs with low plasma von Willebrand factor concentration, although some dogs with these genotypes did not have abnormal bleeding.

Conclusions and Clinical Relevance—Type-1 von Willebrand disease in Doberman Pinschers is associated with the von Willebrand factor gene locus; however, the expression pattern in this breed appears more complex than that of a simple recessive trait. (Am J Vet Res 2001;62:364–369)

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in American Journal of Veterinary Research