Localization and characterization of nucleotide sequences from the canine Y chromosome

We previously reported the identification of a male-specific 658-bp DNA seq uence in dogs. We used a specific primer pair designed for PCR amplificatio n of this fragment with DNA samples from 238 dogs, 6 dingoes and 12 wolves. All 133 male samples amplified the 658-bp sequence, whereas all female sam ples did not. The sequence was not amplified from male DNA samples represen ting other wild canids (jackals, coyotes, foxes). A lambda phage was isolat ed from a canine male genomic library that contained an insert of approxima tely 15 kb of canine genomic DNA, including the male-specific 658-bp sequen ce. This lambda phage was used in fluorescence in-situ hybridization experi ments. It hybridized to the canine Y chromosome together with a lambda clon e containing a segment of the SRY gene and a cosmid clone containing a port ion of the pseudoautosomal region. The male-specific 658-bp sequence was lo cated at the end opposite to the pseudoautosomal region while the SRY gene sequence hybridized near the centromere. Additionally, two (CA)-repeat sequences were identified in the lambda clone that contained the 658-bp sequence. Specific primer pairs were designed to amplify each of the repeats. Primer pair MS34 amplified three different al leles from 13 unrelated canine male DNA samples with a PIC value of 0.40. P rimer pair MS41 amplified five alleles with a PIC value of 0.71. These micr osatellites are the first reported polymorphic sequences in the dog located in the non-recombining portion of the Y chromosome.