Phylogeny of extinct wolves based on nucleotide saguence

本研究旨在对不同机构饲养的已灭绝的日本狼和野狗的骨骼和皮肤标本进行形态观察。我们试图用这些绒毛标本来测定丝裂软骨DNA的部分核苷酸序列。我们可以从荷兰莱顿博物馆保存的标本中获得形态数据和一块皮肤样本。我们还可以从东京国立科学博物馆北海道大学农学系准备皮肤样本。采集了近40个形态数据和9个皮肤样本。多因素分析表明，亚洲大陆的狼与北海道的狼相近，其次是家犬，这些物种与日本狼的亲缘关系较远。以皮肤样本的DNA为模板，以线粒体DNA的细胞色素b基因为引物，聚合酶链式反应可以从5个样本中的2个样本中扩增出DNA片段。我们不能对其他3个样本进行任何DNA扩增。一些淡棕色物质(S)可能会抑制DNA聚合酶。计算机分析表明，一个是未知真核生物的细胞色素b基因，另一个是人类的细胞色素b基因。我们将继续通过各种方法在聚合酶链式反应之前对DNA样本进行纯化。

The present study aimed morphological examination of skeletal and skin specimens of extinct Japanese wolves and wild dogs which have been kept in various institutions. We attempted to determine thenucleotide sequence of part of mitrochonrial DNA using those pelage samples.We could get morphological data and a piece of skin samples from the specimens kept at Leiden museum in the Netherlands. We could also prepare the skin samples from, Faculty of Agriculture of Hokkaido University, National Science Museum, Tokyo. Neary 40 morphological data and 9 skin samples were collected.Multivariate analyzes of the measurement revealed wolves in Asian continent are close to Ezo-wolves in Hokkaido, next is domestic dogs and these species are rather distant relationships with the Japanese wolves.Polymerase chain reaction using DNA from skin samples as template and primer set for cytochrome b gene of mitochondrial DNA could amplify DNA fragment from two specimens out of 5 examined. We could not amplify any DNA for other 3 samples. Some pale brownish substance (s) may inhibit DNA polymerase. Computer analyzes of the sequences revealed that one is cytochrome b gene of unkown eukaryote and the other is cyto-chrome b of human. We will continue to purify the DNA samples before PCR by various methods.