Evaluation of the genetic diversity of Iranian Kurdish horses

The horse is a large land mammal notable for its speed, strength, and endurance. Horses are members of the Equidae family, the horse’s influence on human history and civilization make it one of the most important domestic animals. The horse has various breeds and these breeds can be distinguished from each other based on morphological characteristics (Mahrous et al. 1994). However, this method is not accurate enough. Among molecular markers, microsatellites are suitable for biodiversity evaluation owing to their codominant inheritance, high heterozygosity and distribution across the genome, ease and reliability of scoring. Microsatellites or Short Tandem Repeats (STRs) are used as useful markers found in the DNAs of most species. They are defined as tandem repeats of 2-10 base pair units and are often present as perfect or imperfect repeats. The number of repeats found in any given microsatellite region is sometimes highly variable, with as many as hundreds of copies of the repeat unit in each microsatellite. As a result of their highly polymorphic nature, microsatellites are informative molecular markers that can be applied to research in the fields of population genetics, medical genetics, forensic science, evolutionary biology, and plant breeding. Once potentially useful (i.e. polymorphic) microsatellites are found, PCR primers are constructed from the DNA sequences flanking the microsatellite regions Thus researchers are using microsatellite markers for parentage testing as well as for population genetics studies. In the last decade microsatellite markers have been widely used to assess genetic variability within and between different horse breeds (Nicholas et al. 2009). In the current work genetic diversity of the Kurdish horse breed was studied using 52 individual horses.
Material and Sampling from Kurdish horse was done and their DNAs extracted based on salting out method (Miller et al. 1988). Extracted DNAs was run in an agarose gel and concentration and quality of DNAs was measured by Nano-drop. We used four microsatellite markers that all have been recommended by ISAG for parentage testing. These markers include VHL20, HTG4, AHT4, and HMS7. These loci were amplified by multiplex PCR with fluorescent dye-labeled primers. Polymerase chain reaction was performed using 25 micro liters for each sample and PCR products were separated and analyzed with capillary electrophoresis and the outputs were analyzed using Genmarker software. The results showed there are high genetic diversity within the Kurdish horse population. At VHL20 locus 9 alleles was seen and the most frequent allele at VHL20 locus was 170 bp and the lowest frequent allele was 90 bp also the biggest allele in this locus was 107 bp and the smallest allele was 88 bp, observed heterozygosity in VHL20 locus was 0/75 however the expected heterozygosity was 0/84. At HTG4 locus 8 allele was seen and the most frequent allele at HTG4 locus was 132 bp and the allele with 126 bp had a lowest allelic frequency, the allele with 138bp was the biggest allele at this locus and the smallest allele was 124bp. At this locus observed heterozygosity was 0/73 and expected heterozygosity was 0/72. At AHT4 locus 13 allele was seen also the allele with 156 bp had a highest allelic frequency and the alleles with 147 and 150 bp had a lowest allelic frequency, at this locus the biggest allele was 156bp and the allele with 140 bp was the smallest allele in this locus. Observed and expected heterozygosity at this locus were 0/80 and 0/85 At HMS7 locus 8 alleles was seen and the allele with 183 bp was the most frequent and the allele with 183 bp had a low allelic frequency. At HMS7 locus the biggest allele was 183 bp and smallest allele was 169 bp. Observed and expected heterozygosity was calculated 0/75 and 0/73. In average 9/5 allele per each locus seen in this population. The number of alleles was between 8 and 13. The AHT4 marker had 13 alleles with the highest allelic frequency and the highest heterozygosity. Either of HTG4 and HMS7 markers had 8 alleles and had the lowest allelic frequency and heterozygosity.
Results of this study showed high frequency genetic diversity in Iranian Kurdish population in compare with the other horse breeds. The non-standard mating and mixing of Kurdish horses with other breeds can be the reason of the high diversity.