Genetic characterization of the Russian population of German Classic ponies by STR loci and genes affecting coat colour

The German classic pony is a relatively new breed officially registered in 2000. The German classic pony was created in Germany with the aim of breeding horses that are ideal for children. The ponies are smart, good-tempered and energetic, elegant and handsome, looking like a riding mini-horse. The breed is characterized by presence of attractive silver dapple coat colour of various shades. The silver dapple coat colour in horses is the result of the dominant Silver mutation of the PMEL17 gene that affects the black pigment eumelanin. Typically, the Silver gene changes the black colour of hairs of mane and tail to white, flaxen or grey. German classic ponies have been bred in Russia since 2010. At present, the Russian population of German classic ponies is extremely small. At the beginning of 2023, there were 30 purebred brood mares and 9 stallions. The breeding stock is concentrated on one farm. The unique silver coat colour of the ponies is considered one of the most important traits of selection in the breed. The aim of the study was to evaluate the genetic variability using 17 STR markers and to investigate polymorphism of the 4 genes associated with the coat colours in German classic ponies of the Russian population. A total number of 32 animals were genotyped by using markers of genes MC1R , ASIP , PMEL17 , MATP and 17 STR markers: AHT4, AHT5, ASB2, ASB17, ASB23, CA425, HMS1, HMS2, HMS3, HMS6, HMS7, HTG4, HTG6, HTG7, HTG10, LEX3, VHL20. Hair samples were collected from the German classic ponies of the farm Koshelev V.V. in 2019. DNA was isolated from hair follicles using the commercial reagent kit ExtraGene™ DNA Prep 200 (Laboratory Isogen, Russia). Genotyping for STR markers was performed using the reagent kit for multiplex analysis COrDIS Horse (GORDIZ, Russia). Based on the testing results, the following parameters were calculated: allele frequencies, the number of observed alleles in each locus (A), the effective number of alleles (Ae), expected (He) and observed (Ho) heterozygosity, and the inbreeding coefficient (FIS). The results revealed a relatively high level of genetic variability in the most of studied STR loci. The number of alleles identified across all evaluated STR loci was 94. The number of alleles per locus varied from 3 to 8 with a mean value of 5.53±0.35. The level of the observed heterozygosity (Ho) for 16 autosomal loci was 0.7051±0.0434, the FIS coefficient was -0.0686±0.0196. It was found that the frequencies of the dominant A allele of the ASIP gene and the dominant E allele of the MC1R gene were 0.313 and 0.688, respectively. The frequency of the mutant Silver allele of the PMEL17 gene was 0,563. The mutant Cremello allele of the MATP gene was not found in the group of genotyped ponies. Given four genes that control pigmentation, 14 different genotype variants were identified in the tested animals, indicating a high level of genetic variability in the population. The most common genotypes in the studied population were associated with the most unusual and attractive coat colors.