Polymorphisms Of Prion Protein Alleles In Sheep

The polymorphism of PrP allele is associated with the scrapie and the study on the condition of the PrP alleles in Chinese sheep by the new method is necessary. The current research is to investigate the PrP alleles in sheep by using SSCP and PAGE methods and the results could be used in establishing a sheep herd without the scrapie risk.Total DNA was extracted from peripheral blood samples of 10 sheep. The PrP allele were amplified by polymerase chain reaction (PCR) with a pair primers. The product of PCR was analyzed by single-strand conformation polymorphism (SSCP) and PAGE .The difference of the product was determined. The silver-staining procedure using polyarylamide gel was performed after eletrophoresie.The product of heterozygote and homozygote were purified cloned into PMD18-T Vector, respectly. The clone were selected and amplified by polymerase chain reaction . After digestion with EcolⅠand HindⅢ, the entire clone PrP allele fragment was constructed. The nucleotides were analyzed and amino acid sequences of their homology by DNAstar. Similar methods were used to analyse the PrP allele from 5 sheep.The results indicated that SSCP and PAGE could be used to seek polymorphism PrP allele and PrP is encoded by a single-copy autosomal gene. In this study, it was found that 2 sheep genotype are heterozygote with the phenotypy of 136AA 154RR171HR and other 8 sheep are homozygote with the phenotypy of 136AA 154RR171QQ. It was found that all of 5 sheep are homozygote and 7 polymorphisms in PRNP gene,including 6 previously reported(23L→V,25K→E,136 A→V,154H→R,171Q→R,189 Q→C)and 1 was not reported(24 C→W). VRQ(173)and ARQ(175,133) are associated with susceptibility to scrapie; ARR(7401,201) are associated with resistance to scrapie in several genotype.It could be concluded from the above results that the samples containing the PrP allelic ARQ and VRQ at codons 136,154 and 171 are associated with susceptibility to scrapie 73% in the total. The risk of the occurrence of scrapie in the experimental areas of Chinease sheep was obtained from information of the result.