Simultaneous detection of eight single nucleotide polymorphisms in the ovine prion protein gene

Amino acid polymorphisms in the prion protein gene (PrP) affect the susceptibility of sheep to scrapie, a transmissible spongiform encephalopathy (TSE). In particular, amino acid substitutions at codons 136, 154 and 171 of the ovine PrP gene are associated with different degrees of susceptibility to the classical form of scrapie, caused by 'typical' scrapie strains. Existing genotyping tests for scrapie susceptibility normally interrogate only the single nucleotide polymorphisms (SNPs) most relevant to 'typical' strains. Recently, however, a number of novel variants of the scrapie agent have been discovered. The ability of these new, 'atypical' scrapie variants to infect sheep that are resistant to 'typical' variants has raised concerns about the reduction in genetic variability that may result from intense selection for resistance to classical scrapie. Furthermore, a growing interest in a potential role for specific PrP genotypes in modulating performance traits is also driving a move toward more extensive characterization of haplotypes at the PrP locus. Here, we describe a single-tube method for the interrogation of eight SNPs within seven codons (112, 136, 141, 154, 171, 231 and 241) of the ovine PrP gene. This method is as accurate as sequencing, yet more affordable, and can easily be automated for high-throughput sample screening. Moreover, it can be modified to accommodate genetic variations that are found in local and heritage breeds.