利用微衛星標幟探討臺灣本地山羊之遺傳歧異度

Abstract

臺灣養羊產業為乳品及肉品產業的其他來源之一，且品種保存為種畜生產的重要基礎。然而過去的文獻顯示，在 DNA 層面上臺灣本地山羊的遺傳歧異度尚未研究清楚，又骨形態發生蛋白 15 基因（bone morphogenetic protein 15 gene, BMP15 gene）已證實對許多綿羊及山羊品種為多產基因。臺灣黑山羊品種以多產聞名，本研究利用微衛星標幟探討臺灣黑山羊與其他外國山羊族群的遺傳歧異度，並偵測臺灣本地山羊不同族群 BMP15 基因 exon 2 的多態性與多產之相關。 本研究蒐集來自 6 個山羊品種共 503 頭山羊之血樣（臺灣黑山羊 134、努比亞 185、撒能 46、阿爾拜因 90、波爾 16及吐根堡 32 頭），運用 15 組微衛星標幟進行遺傳變異分析。結果顯示所有的微衛星標幟在所檢測的族群中皆具有多態性，而平均期望雜合度（average expected heterozygosities, HE）與觀測期望雜合度（observed heterozygosities, HO）分別為 0.655±0.026 與 0.602±0.034。不同山羊族群之間的分化程度（FST）為 0.174，範圍從 0.111（SRCRSP001）到 0.292（OarFCB20）。平均期望雜合度於山羊族群的範圍從 0.577±0.034（恒春臺灣黑山羊）到 0.731±0.030（佳里B阿爾拜因），再經由 Fisher 精確檢定後結果顯示所有族群皆符合哈溫平衡（P>0.05）。更進一步地利用鄰近相接法（neighbor-joining）繪製親緣關係樹，結果顯示主要可以分成兩大群集：臺灣黑山羊和其他外國山羊族群，然而波爾族群則另單獨成一分支。而外國山羊族群群集又可分成兩個次群集：努比亞與其他山羊族群（阿爾拜因、撒能與吐根堡族群）群集。此外恆春與花蓮臺灣黑山羊群集有很高的再取樣值（bootstrap value, 96%）。 本研究另蒐集共 6 個品種 447 頭之山羊血樣，並藉由定序分析 BMP15 基因的多態性。結果顯示所有檢測的山羊族群 BMP15 基因皆為 A963/C1050 haplotype，而臺灣本地山羊之平均母羊產羔數（litter size）為 1.93 ± 0.15 頭。同時也在 BMP15 基因exon 2 上第 870 個位置發現新的 C/G 單一核苷酸變異（single nucleotide polymorphism, SNP），C 與 G 交替基因頻率的範圍為 0.52~1.00 到 0.00~0.48，所檢測的山羊族群中皆存在 C/C 基因型，但 G/G 基因型僅存在恆春及花蓮黑山羊族群（25.40 vs. 9.59%）。進一步分析 BMP15 基因新的 SNP 與產羔數的關係，結果顯示在臺灣黑山羊族群 BMP15 基因型與產羔數無顯著差異（P>0.05）。 綜合所述，雜合度的指標（HE, HO, and F-統計值）可供為臺灣本地山羊遺傳歧異度的參考依據，且不同山羊族群之間的遺傳關係與其來源相似。此外，檢測臺灣本地山羊族群之 BMP15 基因 A963/C1050 位置顯示不具有多態性，因此可能需要尋找 BMP15 基因的多態性與多產性的相關。

Goats are one of the important sources of meat and milk in addition to cow in Taiwan. In general, preservation of goat breeds is essential for animal productions. In previous studies, genetic diversity of Taiwan native goats still has not been studied clearly on DNA level. The studies have shown that a major gene, bone morphogenetic protein 15 gene (BMP15), is responsible for prolificacy in goat and sheep. The Taiwan Black is a prolific goat breed in Taiwan. Thus, the objectives of this study were to investigate genetic diversity between Taiwan Black goats and other exotic breeds from different populations based on microsatellite markers, and to detect the polymorphism on the exon 2 of BMP15 gene in populations of Taiwan native goat. In this study, a total of 503 blood samples were collected from six goat breeds (134 Taiwan Black, 185 Nubian, 46 Saanen, 90 Alpine, 16 Boer, and 32 Toggenburg goats), and genotyped using 15 microsatellite markers. The results showed that all microsatellite markers were polymorphic in all the populations in this study. The average expected heterozygosities (HE) and observed heterozygosities (HO) per locus in all populations were 0.655±0.026 and 0.602±0.034, respectively. The average differentiation among populations (FST) was 0.174, and ranged from 0.111 (SRCRSP001) to 0.292 (OarFCB20). The average expected heterozygosities (HE) over all loci varied from 0.577±0.034 (Taiwan Black goats from Hengchun) and 0.731±0.030 (Alpine from Jiali B). The Fisher’s exact test showed that all populations were in Hardy-Weinberg equilibrium (P>0.05). The genetic diversity of Taiwan native goats was analyzed further by neighbor-joining tree. The results suggested that there were mainly two clusters: Taiwan Black goats and others exotic goats, whereas Boer goats were clearly in one branch. There were two subgroups in the cluster of others exotic goats: Nubian goats and the others (Alpine, Saanen, and Toggenburg). Furthermore, the cluster of Taiwan Black goats from Hengchun and Hualien had high bootstrap value (96%). In BMP15 gene, a total of 447 blood samples were collected from six goat breeds and genotyped by DNA sequencing. The results showed that all the tested goats had A963/C1050 haplotype in BMP15 gene. The average litter size of all the tested does was 1.93 ± 0.15 heads. Besides, our results found that there was a novel C/G single nucleotide polymorphism (SNP) in BMP15 gene exon 2 at the 870 site. The ranges of C and G allele frequencies were 0.52~1.00 and 0.00~0.48, respectively. The C/C genotype was found in all the tested populations, but the G/G genotype was only in Taiwan Black goat populations from Hengchun and Hualien (25.40 vs. 9.59%). There was not significance difference between the BMP15 genotypes and litter size (P>0.05) in the novel SNP of BMP15 gene. In conclusion, the values of heterozygosities (HE, HO, and F-statistics) suggested that Taiwan native goats were a reservoir of goat diversity. Relationships between populations agreed with what know about their origin. In addition, the results of polymorphism at A963/C1050 site in BMP15 gene indicated that none of polymorphism in all Taiwan native goat breeds, so it is necessary to further study about relationship between BMP15 gene polymorphic and the prolificacy in Taiwan.