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Scrapie is a fatal, neurodegenerative disease occurring in sheep, goats and mufflons. It is commonly believed that its infectious agent is a protease-resistant form of host-encoded prion protein (PrP). Polymorphism of the coding region of PrP genes has been analyzed in many countries. Polymorphism of codons 136, 154 and 171 has been associated with outbreaks of scrapie. Valine (V) in codon 136, arginine (R) in codon 154, glutamine (Q) and histidine (H) in codon 171 are associated with susceptibility to scrapie while histidine (H) in codon 154 and arginine (R) in codon 171 are associated with resistance to the illness. Alanine (A) in codon 136 is associated with low susceptibility to scrapie.
Short tandem repeat (STR) loci, i.e. microsatellites are a class of genetic markers commonly used for population studies and parentage control. This study determined the usefulness of microsatellite markers recommended by International Society for Animal Genetics (ISAG) for identification and pedigree analysis in horses based on the example of Polish Hucul horse population (Equus caballus). The set of seventeen microsatellites loci was tested (AHT4, AHT5, ASB2, HMS2, HMS3, HMS6, HMS7, HTG10, HTG4, HTG6, HTG7, VHL20, ASB17, ASB23, CA425, HMS1, LEX3) for 216 individuals. All samples were genotyped and mean number of alleles per locus was estimated (7.00). Means of observed (Ho) and expected (He) heterozygosity were calculated 0.7288 and 0.7027, respectively. The observed heterozygosity was similar to the results of research on Hucul horse population in another area of Carpathians Mountains. The average polymorphism information content (PIC) for analyses of seventeen microsatellite markers indicates the usefulness of this set of markers for Hucul horse parentage testing.
The paper describes adaptation of primer extension method for genotyping scrapie-associated codons 136, 141 and 156 in sheep PRNP gene. As the method failed to genotype the polymorphism in codon 171 in some samples, this polymorphism was excluded from the assay. Such genotyping problems were not observed in earlier studies describing the method and cannot be easily explained. However, full accordance of genotypes in codons 136, 141 and 156 with sequencing results was obtained and thus the method can be recommended for routine application. Simultaneously, we strongly recommend to pay special attention when using the minisequencing method for genotyping codon 171 in sheep PRNP.
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