BRIEF NOTE
The domestic horse harbours Y-chromosomal microsatellite polymorphism only on two widely distributed male lineages Nina Kreutzmann*†, Gottfried Brem* and Barbara Wallner* *Department of Biomedical Sciences, Institute of Animal Breeding and Genetics, University of Veterinary Medicine, Veterinaerplatz 1, 1210, Vienna, Austria; †Clinic of Small Animal Medicine, LMU University of Munich, Geschwister-Scholl-Platz 1, 80539, Munich, Germany Accepted for publication 16 February 2014
Source/description: Mitochondrial DNA (mtDNA) and the paternally inherited portion of the Y chromosome (NRY) are inherited uniparentally and do not recombine. They provide a tool for constructing female and male genealogical histories and have therefore been widely analysed in a broad range of wild and domesticated animals. In livestock, mtDNA and NRY markers act as an informative tool to elucidate the origin of breeds and to trace maleand female-mediated gene flow. Paradoxically, horses (Equus caballus) have the largest mtDNA diversity among domestic species, and mtDNA has been widely used to investigate the origin of domestic horse breeds, but only limited sequence diversity has been described on the NRY.1,2 In a recent paper, we presented the first polymorphic Y-chromosomal markers for paternal lineage analysis in modern domestic horses. They resulted in only six haplotypes, all clearly distinct from the Przewalski horse (Equus przewalskii).3 Only single mutational steps separate the modern horse Y-chromosomal haplotypes from each other. Horse paternal lineages seem therefore closely related, but Y-chromosomal diversity might be underestimated on the sequence level due to ascertainment bias.4 Highly mutable microsatellite markers may help to increase the resolution, as shown in cattle.5 An initial investigation of five Y-chromosomal microsatellites revealed no polymorphism and thus supported a very close relationship and recent origin of the modern horse haplotypes.3 In this study, we investigate Y-chromosomal lineages in greater depth using eight newly identified Y-chromosomal microsatellite markers in a comprehensive sample set containing all modern and Przewalski horse Y-chromosomal haplotypes. Isolation of microsatellite loci and assessment of genetic diversity:
We identified eight di-, tri- or tetra-nucleotide microsatellites within Y-chromosomal BAC-derived reference contigs (Table S1). Primers flanking the microsatellites were designed using PRIMER3 software (Whitehead Institute for Biomedical Research), and Y-specificity was checked by comparative amplification of genomic DNA from male and
© 2014 Stichting International Foundation for Animal Genetics
doi: 10.1111/age.12149 female horses (Fig. S1). DNA samples were collected from 104 male purebred domestic horses from 41 different breeds and three Przewalski horses. They were screened for their Y-chromosomal haplotype (as described in3) and also for microsatellite polymorphism (as described in6 and Table S1). A detailed list of the horses screened and their Y-chromosomal haplotype is given in Table S2. Comments: Most Y chromosomes showed no microsatellite
polymorphism in domestic horses. Only three, apparently unrelated, horses harboured mutant microsatellite alleles: EcaYP9 in a Hucul and a Mongolian horse on different backbones (HT2 and HT1 respectively) and EcaYNO4 in a Shetland pony on HT1. Although the microsatellite phenotype is identical in the Hucul and the Mongolian horse, the mutation must have arisen independently due to the different backbone. We therefore postulate three recent independent mutations. The detection of two paternal lines in the Przewalski horse confirms previous findings.3,6 Our data reinforce previous findings of a severe reduction of the genetic variability on the horse Y chromosome and provide further evidence for a very recent origin of modern horse paternal lineages. In contrast, the average number of alleles is about seven to eight per microsatellite when screening randomly chosen autosomal microsatellites in modern horses.7 We note that all three Y-chromosomal microsatellite mutations occurred on the backbone of the two major lines HT1 and HT2. This suggests that the derived haplotypes found in Northern horses (HT4, HT5, HT6)3 originated relatively recently compared to the common haplotypes HT1 and HT2. Conflict of interest: The authors declare no conflict of interest. References 1 Achilli A. (2012) Proc Natl Acad Sci USA 109, 2449–54. 2 Lindgren G. (2004) Nat Genet 36, 335–6. 3 Wallner B. (2013) PLoS One 8, e60015. 4 Lenstra JA. (2012) Anim Genet 43, 483–502. 5 Edwards C. (2011) PLoS One 6, e15922. 6 Wallner B. (2004) J Hered 95, 158–64. 7 Mittmann E. (2010) J Hered 101, 246–50 Correspondence: B. Wallner ([email protected])
Supporting information Additional supporting information may be found in the online version of this article. Figure S1. Y-specificity of newly identified microsatellite markers. Table S1. Microsatellite markers developed in this study. Table S2. Haplotype screening results.
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The domestic horse harbours Y-chromosomal microsatellite polymorphism only on two widely distributed male lineages Nina Kreutzmann*†, Gottfried Brem* and Barbara Wallner* *Department of Biomedical Sciences, Institute of Animal Breeding and Genetics, University of Veterinary Medicine, Veterinaerplatz 1, 1210, Vienna, Austria; †Clinic of Small Animal Medicine, LMU University of Munich, Geschwister-Scholl-Platz 1, 80539, Munich, Germany Accepted for publication 16 February 2014
Source/description: Mitochondrial DNA (mtDNA) and the paternally inherited portion of the Y chromosome (NRY) are inherited uniparentally and do not recombine. They provide a tool for constructing female and male genealogical histories and have therefore been widely analysed in a broad range of wild and domesticated animals. In livestock, mtDNA and NRY markers act as an informative tool to elucidate the origin of breeds and to trace maleand female-mediated gene flow. Paradoxically, horses (Equus caballus) have the largest mtDNA diversity among domestic species, and mtDNA has been widely used to investigate the origin of domestic horse breeds, but only limited sequence diversity has been described on the NRY.1,2 In a recent paper, we presented the first polymorphic Y-chromosomal markers for paternal lineage analysis in modern domestic horses. They resulted in only six haplotypes, all clearly distinct from the Przewalski horse (Equus przewalskii).3 Only single mutational steps separate the modern horse Y-chromosomal haplotypes from each other. Horse paternal lineages seem therefore closely related, but Y-chromosomal diversity might be underestimated on the sequence level due to ascertainment bias.4 Highly mutable microsatellite markers may help to increase the resolution, as shown in cattle.5 An initial investigation of five Y-chromosomal microsatellites revealed no polymorphism and thus supported a very close relationship and recent origin of the modern horse haplotypes.3 In this study, we investigate Y-chromosomal lineages in greater depth using eight newly identified Y-chromosomal microsatellite markers in a comprehensive sample set containing all modern and Przewalski horse Y-chromosomal haplotypes. Isolation of microsatellite loci and assessment of genetic diversity:
We identified eight di-, tri- or tetra-nucleotide microsatellites within Y-chromosomal BAC-derived reference contigs (Table S1). Primers flanking the microsatellites were designed using PRIMER3 software (Whitehead Institute for Biomedical Research), and Y-specificity was checked by comparative amplification of genomic DNA from male and
© 2014 Stichting International Foundation for Animal Genetics
doi: 10.1111/age.12149 female horses (Fig. S1). DNA samples were collected from 104 male purebred domestic horses from 41 different breeds and three Przewalski horses. They were screened for their Y-chromosomal haplotype (as described in3) and also for microsatellite polymorphism (as described in6 and Table S1). A detailed list of the horses screened and their Y-chromosomal haplotype is given in Table S2. Comments: Most Y chromosomes showed no microsatellite
polymorphism in domestic horses. Only three, apparently unrelated, horses harboured mutant microsatellite alleles: EcaYP9 in a Hucul and a Mongolian horse on different backbones (HT2 and HT1 respectively) and EcaYNO4 in a Shetland pony on HT1. Although the microsatellite phenotype is identical in the Hucul and the Mongolian horse, the mutation must have arisen independently due to the different backbone. We therefore postulate three recent independent mutations. The detection of two paternal lines in the Przewalski horse confirms previous findings.3,6 Our data reinforce previous findings of a severe reduction of the genetic variability on the horse Y chromosome and provide further evidence for a very recent origin of modern horse paternal lineages. In contrast, the average number of alleles is about seven to eight per microsatellite when screening randomly chosen autosomal microsatellites in modern horses.7 We note that all three Y-chromosomal microsatellite mutations occurred on the backbone of the two major lines HT1 and HT2. This suggests that the derived haplotypes found in Northern horses (HT4, HT5, HT6)3 originated relatively recently compared to the common haplotypes HT1 and HT2. Conflict of interest: The authors declare no conflict of interest. References 1 Achilli A. (2012) Proc Natl Acad Sci USA 109, 2449–54. 2 Lindgren G. (2004) Nat Genet 36, 335–6. 3 Wallner B. (2013) PLoS One 8, e60015. 4 Lenstra JA. (2012) Anim Genet 43, 483–502. 5 Edwards C. (2011) PLoS One 6, e15922. 6 Wallner B. (2004) J Hered 95, 158–64. 7 Mittmann E. (2010) J Hered 101, 246–50 Correspondence: B. Wallner ([email protected])
Supporting information Additional supporting information may be found in the online version of this article. Figure S1. Y-specificity of newly identified microsatellite markers. Table S1. Microsatellite markers developed in this study. Table S2. Haplotype screening results.
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