제주마 모색 분포 특성과 얼루기 모색의 분자유전학적 연구

Abstract

Jeju horses are the only horse genetic resource native to South Korea. The present study was conducted to determine the color of Jeju horses' coats and to ensure a basic data of coat color diversity by investigating the coat color characteristics and the molecular genetic characteristics of Jeju horses with white patch. To characteristics the coat color of Jeju horses, the genotypes of MC1R and ASIP genes that are related to the basic black, bay, and chestnut coat colors of 1,465 heads of registered Jeju horses, 119 heads of thoroughbred, and 228 heads of crossbred(Jeju horse × thoroughbred) were analyzed. In addition, the genotypes of STX17 and ECA3-inversion that are related to gray and white coat colors were also studied. It was found that, the frequencies of genotypes E+/E+, E+/Ee, and Ee/Ee of the MC1R gene were identified 0.122, 0.447, and 0.429, respectively. Although thoroughbred and crossbred horses showed similar frequencies to those of Jeju horses, however thoroughbred horses has shown a lower frequency of genotype Ee/Ee. Among Jeju horses, the frequencies of the genotypes AA/AA, AA/Aa, and Aa/Aa of the ASIP gene were found at 0.46, 0.448, and 0.091, respectively. The black coat color is expressed by the combination of genotype E+/- of the MC1R gene and the genotype Aa/Aa of the ASIP gene. Because the frequency of the genotype Aa/Aa among Jeju horses was shown to be 0.091, the rate of expression of black coat color is estimated to be 5.1% or lower. Among three groups of horses, Jeju horses, crossbred horses, and thoroughbred horses, the frequencies of haplotype were found E+-AaAa which indicates black coat color were shown to be 0.042, 0.079, and 0.025, respectively, the highest frequency among of the crossbred horses. The frequency of bay coat color was the highest among thoroughbred horses, and Jeju horses and crossbred horses showed similar frequencies. The frequencies of genotypes To/To and To/+ that are expressed as phenotype Wolla (white patch) among Jeju horses were shown 0.1% and 11.9% respectively. Therefor 12% of Wolla coat color frequency stipulated among Jeju horses. The genotype To/To was not identified in crossbred horses, while the frequency of the genotype To/+ was 3.9%, which is lower frequency of Wolla coat color compared to Jeju horses. On the other hand, the allele did not appear in the thoroughbred horses. The frequencies of genotype G/g for the expression of gray coat color among thoroughbred and crossbred horses were shown to be low at 6.7% and 5.2%, respectively. On the counterpart, the homozygote frequency and heterozygote frequency of G allele among Jeju horses was shown to be 0.2% and 68%, respectively, indicating that gray coat color is prevalent among Jeju horses. The genes MC1R, ASIP, and ECA3-inversion related with coat color maintained by Hardy-Weinberg equilibrium, however STX17 in Jeju horses was not in the equilibrium state (p<0.001). It was thought to be responsible by selection of a gray coat color stallion for the maintenance of a group of Jeju horses The allele and genotype frequencies of the MC1R and ASIP genes of Jeju horses have not changed very much with each generation. The frequency of the Wolla To allele decreased gradually from 0.126 to 0.034 within 0 to 2.5 generation. Thus it was assumed that the occurrence of the Wolla coat color in Jeju horses significantly decreased with each generation (p<0.05). The frequency of the G allele of the STX17 gene that expresses gray significantly increased from 0.192 in generation 0 to 0.386 in generation 2.5 (p<0.05). In addition, the frequency of the genotype G/g of STX17 gene that expresses gray also increased by two times from 0.384 to 0.764 at 0 to 2.5 generation, indicating that the frequencies of allele and genotype that express gray rapidly increased with successive generations. The genetic classification of Jeju horses' Wolla coat color was conducted using molecular genetic analysis methods to detect the presence of the tobiano, overo, and sabino genes. It was identified that in the case of Jeju horses' Wolla coat color, all ECA3-inversions that are related to the tobiano gene has the To allele, while all EDNRB related to the overo gene and the KIT intron 16 SNP related to the sabino gene were wild types. Therefore, Jeju horses' Wolla coat color is thought to be due to the tobiano gene. According to the results of a genome-wide association study (GWAS) analysis of horses' white patch, 72 SNPs in the equine genome satisfied the significance level of p<0.001. The significant SNPs were distributed widely from 21 Mbp to 85.1 Mbp in chromosome number 3. The most significant SNP was located at 48853535bp (BIEC2-819509), and its p value was shown to be 1.01× 10-22. To identify related genes in SNP loci significantly related to horses' white patch, regional plots were prepared. The GPRIN3 gene and the TIGD2 gene could be identified in the vicinity of the most significant SNP locus, and MC1R and KIT genes were also shown to exist in significant regions. The GPRIN3 gene's DNA sequence was analyzed and six SNPs were identified; no insertion/deletion mutation was found. Among the identified SNPs, c.1114G>A and c.1476G>A were identified as being synonymous, and c.1226C>T, c.1388C>T, and c.1790G>A were identified as substituting amino acid sequences with p.372V>I (Val>Ile), p.409S>L (Ser>Leu), and p.463T>M (Thr>Met), respectively. The correlation between SNPs and white patch in the GPRIN3 gene was analyzed, and the result showed high LOD values in the range of 60.9–121.2. Four SNPs were identified in the TIGD2 gene, and in the case of c.*152G>A, differences between solid and white patch base sequences could be identified. The correlation between TIGD2 c.*152G>A SNP and white patch was analyzed, and the result showed a very high LOD value of 206.6. The present study shows that the diversity of Jeju horse coat colors has been decreasing. In particular, gray color-related gene frequencies were shown to be high to account for most coat colors, and the frequencies significantly increased with successive generations. Therefore, to maintain the diversity of Jeju horses' coat colors, coat color factors should be reviewed in the stage of selecting stallions. The spotted coat color appearing in Jeju horses was identified as genetically due to the tobiano gene. The third chromosome of horses was shown to be highly correlated with spotted coat color, and TIGD2 c.*152G>A, GPRIN3 c.1114G>A, GPRIN3 c.*331A>G, and BIEC-819509 SNP are considered assistant markers to identify tobiano coat color.