Identification of a 6 base pair insertion in West Highland White Terriers with erythrocyte pyruvate kinase deficiency

Barbara J. Skelly From the Section of Medical Genetics, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, PA 19104-6010.

Search for other papers by Barbara J. Skelly in
Current site
Google Scholar
PubMed
Close
 Vet MB, PhD
,
Mary Wallace From the Section of Medical Genetics, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, PA 19104-6010.

Search for other papers by Mary Wallace in
Current site
Google Scholar
PubMed
Close
 VMD
,
Yashoda R. Rajpurohit From the Section of Medical Genetics, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, PA 19104-6010.

Search for other papers by Yashoda R. Rajpurohit in
Current site
Google Scholar
PubMed
Close
 BS
,
Ping Wang From the Section of Medical Genetics, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, PA 19104-6010.

Search for other papers by Ping Wang in
Current site
Google Scholar
PubMed
Close
 MS
, and
Urs Giger From the Section of Medical Genetics, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, PA 19104-6010.

Search for other papers by Urs Giger in
Current site
Google Scholar
PubMed
Close
 Dr med vet

Click on author name to view affiliation information

Abstract

Objective

To define the disease-causing mutation in West Highland White Terriers (WHWT) with erythrocyte pyruvate kinase (R-PK) deficiency and to design a genetic test capable of recognizing affected (homozygous) and carrier (heterozygous) dogs.

Animals

3 anemic WHWT littermates and 1 unaffected littermate; 16 dogs from the same kennel, including 4 unrelated, phenotypically normal dogs (control dogs), and 12 for which PK activity was not known; 2 PK-deficient Basenjis; 2 PK-deficient Beagles; 4 unaffected English Springer Spaniels; and 1 mixed-breed dog.

Procedures

cDNA was cloned and sequenced, and cDNA sequences were compared with the published sequence for canine R-PK cDNA to identify the putative disease-causing mutation. Genomic DNA spanning the affected region was cloned and sequenced to verify the mutation. Subsequently, polymerase chain reaction primers were designed to amplify the section of the gene containing the mutation from DNA in blood or buccal swab samples. Gel electrophoresis allowed assignment of genotypes on the basis of allele separation.

Results

4 single base polymorphisms attributable to sequencing errors in the published sequence were identified, along with a 6 base pair (bp) insertion in exon 10 that was recognized as a putative disease-causing mutation. An identical insertion was found in genomic DNA. Amplification of genomic DNA yielded a 117 bp product for genotypically normal dogs and a 123 bp product for WHWT homozygous for PK deficiency. Carriers had 1 copy of each allele and variable heteroduplex structures.

Conclusions and Clinical Relevance

A 6 bp insertion in the C domain of R-PK was identified in whwt with PK deficiency. Affected and carrier dogs could be distinguished with a genetic test. (Am J Vet Res 1999;60:1169-1172)

Abstract

Objective

To define the disease-causing mutation in West Highland White Terriers (WHWT) with erythrocyte pyruvate kinase (R-PK) deficiency and to design a genetic test capable of recognizing affected (homozygous) and carrier (heterozygous) dogs.

Animals

3 anemic WHWT littermates and 1 unaffected littermate; 16 dogs from the same kennel, including 4 unrelated, phenotypically normal dogs (control dogs), and 12 for which PK activity was not known; 2 PK-deficient Basenjis; 2 PK-deficient Beagles; 4 unaffected English Springer Spaniels; and 1 mixed-breed dog.

Procedures

cDNA was cloned and sequenced, and cDNA sequences were compared with the published sequence for canine R-PK cDNA to identify the putative disease-causing mutation. Genomic DNA spanning the affected region was cloned and sequenced to verify the mutation. Subsequently, polymerase chain reaction primers were designed to amplify the section of the gene containing the mutation from DNA in blood or buccal swab samples. Gel electrophoresis allowed assignment of genotypes on the basis of allele separation.

Results

4 single base polymorphisms attributable to sequencing errors in the published sequence were identified, along with a 6 base pair (bp) insertion in exon 10 that was recognized as a putative disease-causing mutation. An identical insertion was found in genomic DNA. Amplification of genomic DNA yielded a 117 bp product for genotypically normal dogs and a 123 bp product for WHWT homozygous for PK deficiency. Carriers had 1 copy of each allele and variable heteroduplex structures.

Conclusions and Clinical Relevance

A 6 bp insertion in the C domain of R-PK was identified in whwt with PK deficiency. Affected and carrier dogs could be distinguished with a genetic test. (Am J Vet Res 1999;60:1169-1172)

All Time Past Year Past 30 Days
Abstract Views 0 0 0
Full Text Views 27 27 2
PDF Downloads 29 29 1
Advertisement