Untersuchungen zu familiären und rassespezifischen ...

alternatively spliced variants of SILV differing by the presence or absence of seven
aa. 9 An additional splice removes a 126 bp exon and results in two short isoforms of
626 and 619 aa. 10 Si (silver, melanocyte protein 17, pmel17, gp100, NC_000076.4) is
involved in premelanosome biogenesis and its expression is almost exclusive to the
skin and eyes. 11 Si caused hair colour dilution in an inbred strain of black mice over
time. 11
In Dalmatian dogs, CCSD was linked to markers on CFA10, near the merle
locus (K. Mieskes & O. Distl, unpublished results). Therefore, the SILV gene was
examined in this study as a candidate for blue eye colour and CCSD in Dalmatian
dogs.
Animals: We collected samples of 88 unrelated purebred Dalmatian dogs and
classified these dogs into four groups, each with 22 animals. The first group
contained bilaterally hearing dogs with brown eyes, the second group was composed
of uni- or bilaterally deaf dogs with brown eyes, the dogs of the third group were
bilaterally hearing with at least one totally or partially blue eye, and group four
consisted of uni- or bilaterally deaf dogs with totally or partially blue eyes. The
hearing status of these dogs was diagnosed by veterinarians using BAER (brain stem
auditory evoked response) tests. Furthermore, blood samples were available from a
black Labrador retriever, a red merle Australian shepherd and two double merle Old
German sheepdogs.
Sequence and mutation analysis: Genomic DNA was extracted from EDTA-blood
samples of the 88 Dalmatian dogs and the four dogs of other breeds including a
Labrador retriever, an Australian shepherd and two Old German sheepdogs. The
intron 10/exon 11 boundary of SILV carrying the insertion mutation was PCR
amplified using the same conditions previously described (Table S1) 7 and separated
by agarose gel electrophoresis. In order to find out whether mutations other than the
SINE insertion in SILV could play a role in CCSD, all eleven SILV exons with their
flanking sequences were PCR amplified, sequenced and used for mutation analysis
in five Dalmatian dogs with different degrees of pigmentation and hearing status.
These dogs included one bilaterally hearing with intense pigmentation (brown eyes