Genetic structure of the local Yakutian horse population for genes MC1R, ASIP, DMRT3, and MSTN

The Yakutian horse is believed to be one of the oldest breeds. The breed has unique morphological characteristics and is well adapted to survive within the Arctic Circle. Yakutian horses have compact body conformation and extremely thick winter coats with long mane and tail. In the Yakutian breed dominate light coat colours: gray and dun. The gray and dun coat colours of Yakutian horses are their natural camouflage. The Yakutian horse is multipurpose breed, because the local horses have been used by people not only for the production of milk and meat, but also as transport animals. In this paper, the genetic structure of the native Yakutian breed was characterized using markers of four genes that are associated with important selected traits in different modern populations of domestic horses (Equus caballus). The aim of our study was to investigate the polymorphism of the ASIP and MC1R genes that determine skin and hair pigmentation, as well as to assess the occurrence of mutations in the MSTN (g.66493737C>T) and DMRT3 (g.22999655C>A) genes associated with athletic performance and locomotion in domestic horses. Hair samples were collected from 45 adult purebred Yakutian horses ( Equus caballus ), including 11 samples from animals of the indigenous type and 34 samples from animals of the Yana type. DNA was isolated using ExtraGene™ DNA Prep 200 reagents (Isogen Laboratory, Russia). Genotyping for the SNP marker C>T of the MC1R gene was carried out using the PCR-RFLP (PCR-restriction fragment length polymorphism) method according to L. Mark-lund et al. (1996). Detection of 11 bp deletion in the ASIP gene was carried out according to the method described by S. Rieder et al. (2001). Allele nomenclature was used according to M. Reißmann (2009): E - dominant wild-type allele, e - recessive (mutant) allele ( MC1R ); A - dominant wild-type allele, a - recessive (mutant) allele ( ASIP ). The SNP mutation in the MSTN gene (g.66493737C>T) was detected by the amplification-created restriction site-PCR (ACRS-PCR) method described by M. Gábor et al. (2014). Genotyping of DNA samples for the SNP marker of the DMRT3 gene (g.22999655C>A) was performed by PCR-RFLP method, C>A polymorphism was detected using restriction endonuclease HpyF3I (Thermo Scientific, Lithuania). Frequencies of alleles, frequencies of genotypes in the population and observed heterozygosity were calculated. Polymorphism of the ASIP and MC1R genes observed in Yakutian horses demonstrated a predominance of allelic variants that determine the synthesis of eumelanin, the darker type of the pigment. In the studied group of horses the frequency of the dominant E allele of the MC1R gene that determines the production of the black pigment eumelanin, was 0.711. The number of homozygous carriers of the recessive mutation of the MC1R gene ( e allele) that determines production of red pigment pheomelanin was 13.3 %. The frequency of the dominant A allele of the ASIP gene that limits the synthesis of the black pigment eumelanin and affects the character of its distribution was 0.400. The number of homozygous carriers of the recessive mutation of the ASIP gene ( a allele) among the tested Yakutian horses was 40 %. This is relatively high value, because in the most of modern horse breeds, the recessive a allele of the ASIP gene is rather rare. In total, eight different genotypes were identified for two key genes affecting skin and hair pigmentation. The most typical genotypes for Yakutian horses were E/E-A/a and E/E-a/a . The character of skin and hair pigmentation in the Yakutian horses could have an adaptive meaning for survival within the Arctic Circle. The frequency of the mutant variants of genes DMRT3 (g.22999655C>A) and MSTN (g.66493737C>T) in the tested horses were 0.011 and 0.022, respectively. Obviously, being presented in the population at a low frequency, the mutant variants of the DMRT3 and MSTN genes have no selection value, because historically, the Yakutian horse has served people as a transport animal in the forest and swampy areas, where only riding is suitable and the most convenient gait is walk.