タイモウ カラ チュウシュツ シタ DNA ヲ モチイタ オオアシトガリネズミ Sorex unguiculatus ノ セイ ハンベツ

Abstract

オオアシトガリネズミは外見による性判別が難しい上に,保定による外部生殖器の観察が野生由来の飼育個体に多大なストレスを与える可能性もある。そこで本研究では,同種の体毛を材料としたPCRによる性判別法について検討した。毛による性判別に先立ち,解剖して生殖腺で性別を確認した個体から肝臓を採取し,肝臓由来のゲノムDNAで性判別を行った。雄特異的プライマーはヨーロッパトガリネズミで雄特異的産物の検出が報告されているDBY8,雌雄共通プライマーはスンクス(ジャコウネズミ)におけるAqp3の相補的DNA配列からデザインしたAQP3を用いた。その結果,DBY8では雄,AQP3では雌雄の両者において明瞭な1本バンドのPCR産物が検出され,それらによる性判別の結果は,解剖で確認した性別と一致した。次いで体毛に由来するゲノムDNAで性判別を行ったところ,解剖で確認した性別と完全に一致した。以上より,生体から反復して簡便に採取できる体毛を材料として,PCRによるオオアシトガリネズミの性判別法を確立した。Sexing of the long-clawed shrew based on its external features is relatively difficult. And physical restraint to observe their external genitalia often causes extreme levels of stress, because the wild shrews are unaccustomed to handling. In this study, we employed polymerase chain reaction (PCR) analysis of genomic DNA (gDNA) from the hair of the shrews as an alternative method for sexing. In the preliminary experiment, the sexes of the shrews were confirmed by observing their gonads after dissection, prior to PCR analysis using gDNA extracted from the livers. DBY8, a male-specific primer set for the European shrew, was utilized as the male-specific primer set. AQP3, designed from the complementary DNA sequence of Aqp3 from musk shrew, was utilized as the gender-neutral primer set. A single, clear PCR product was detected in the gDNA from males by using DBY8 and from both sexes by using AQP3 after electrophoresis. The results of sexing by PCR analysis of the hair gDNA were completely consistent with the sexing as confirmed by observing the dissected gonads. In conclusion, we developed a reliable method of sexing by using PCR analysis of gDNA extracted from the hair of the long-clawed shrew, which is a specimen that can be collected non-invasively and repeatedly