Successful Molecular Cloning and Nucleotide Sequence Determination of Partial Amelogenin (AMELX) Exon DNA Fragment Recovered from a Mounted Taxidermic Pelt Specimen Tentatively Identified as an Extinct Wolf Species, Canis lupus hodophilax Temminck, the Japanese Wolf and Stocked at School of Agriculture and Life Sciences, the University of Tokyo

Abstract

The Japanese wolf, Canis lupus hodophilax Temminck, which putatively became extinct decades ago, remains a taxonomical or phylogenetic enigma. In Japan, 3 mounted taxidermic pelt specimens believed to represent C. lupus hodophilax have been stocked at the University of Tokyo, the National Science Museum and Wakayama University, respectively. Using an improved method developed by our group for the extraction and the PCR amplification of ancient DNA from preserved mammalian pelts, a 351-nucleotide partial genomic DNA stretch of the X-linked gene coding for amelogenin (AMELX) was successfully cloned from a skin specimen derived from the mounted female taxidermic pelt specimen of the University of Tokyo, and its nucleotide sequence was determined. Two allelic lupine AMELX sequences, i. e., the J-type (accession no. AB080688, submitted to DDBJ on 1/Mar./2002) and the R-type (accession no. AB080689, submitted to DDBJ on 1/Mar./2002) were identified. Both the J-type and the R-type alleles differed considerably at the nucleotide as well as the amino acid level from the corresponding AMELX region shared by the dog and the Mongolian wolf. Although the presently discovered lupine variants of the AMELX partial sequence are of great interest from the point of view of taxonomical identity of the Japanese wolf, our preliminary results (unpublished) strongly indicate that polymorphism in the nucleotide as well as the amino acid sequence is likely to exist in this particular region of AMELX among different breeds of domestic dogs, Canis familiaris. Further molecular analysis of the intraspecific as well as the interspecific variations in the AMELX DNA will be needed to gain clear insight into the taxonomical and phylogenetic positions of the Japanese wolf.