Real-time detection of the mutation responsible for progressive rod-cone degeneration in Labrador Retriever dogs using locked nucleic acid TaqMan probes.
Progressive rod-cone degeneration (prcd) is a late onset, autosomal recessive, inherited disease in dogs caused by a G > A substitution in the PRCD locus. prcd has been reported in more than 18 breeds, including Labrador Retriever dogs. In this study, a real-time polymerase chain reaction (PCR) assay, exploiting the features of locked nucleic acid (LNA) fluorescent-labeled probes, was developed to genotype the sequence variants responsible for the disease. Two Labrador Retrievers were diagnosed with prcd by ophthalmological examination performed by a panelist of the Italian hereditary eye disease control program. The 2 dogs, as well as 8 related and 14 unrelated Labrador Retrievers, were genotyped with both direct sequencing of the disease locus and real-time LNA TaqMan PCR assay. Even though the region surrounding the mutation was predicted to be highly structured, making probe annealing difficult, the real-time PCR assay allowed researchers to correctly genotype the dogs in all cases with a sensitivity threshold of 4 ng/reaction of genomic DNA. A real-time PCR assay will allow a high-throughput analysis of a larger cohort of dogs, thereby enabling researchers to investigate the prevalence of the mutated allele in the affected breeds.
Gentilini
Fabio
F
Veterinary Clinical Department, Alma Mater Studiorum University of Bologna, Via Tolara di Sopra 50, 40064 Ozzano dell'Emilia, Bologna, Italy. fabio.gentilini@unibo.it
Rovesti
Gian Luca
GL
Turba
Maria Elena
ME
D000818:Animals; D001284:Atrophy; D001766:Blindness / Q000235:genetics / Q000662:veterinary; D002874:Chromosome Mapping; D002886:Chromosomes, Human, Pair 17; D017931:DNA Primers; D004285:Dogs / Q000235:genetics*; D005136:Eye Proteins / Q000235:genetics; D005838:Genotype; D006801:Humans; D009154:Mutation; D016133:Polymerase Chain Reaction / Q000379:methods; D020641:Polymorphism, Single Nucleotide; D012160:Retina / Q000473:pathology*; D017949:Retinal Cone Photoreceptor Cells / Q000473:pathology*; D012162:Retinal Degeneration / Q000235:genetics* / Q000473:pathology / Q000662:veterinary*; D017948:Retinal Rod Photoreceptor Cells / Q000473:pathology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity
D017931:DNA Primers; D005136:Eye Proteins