Technical Abstract: The ruminant transmissible spongiform encephalopathies (TSEs) are a heterogeneous group of fatal neurodegenerative disorders affecting cattle, sheep, goats, deer and elk. The TSE of mule deer, chronic wasting disease (CWD), was reported in the 1980s in a small region of Colorado. The disease is now reported in white tailed deer and elk in several areas of the US and Canada. CWD is the subject of aggressive control programs, including depopulation of affected captive and free ranging groups. Ovine scrapie, the related TSE of sheep, is controlled by host genetics. Polymorphisms in the PRNP gene are associated with relative disease resistance in many breeds of sheep and scrapie losses can be prevented through selective breeding programs. Under field conditions, alleles associated with relative susceptibility are identified by DNA sequence analysis of the open reading frame of the PRNP gene in affected animals. In this study, sequence analysis of the PRNP gene from mule deer showed polymorphisms at specific sites in the gene from more than 99% of the animals tested. Individual deer were shown to have more than 2 alleles, demonstrating that the gene had been duplicated. A bacterial artificial chromosome library of mule deer DNA was constructed and screened for exon 1 of the PRNP gene. Analysis of the clones revealed a full length functional gene and a processed pseudogene with characteristics of previously described retrotransposons. cDNA analysis indicated that the pseudogene was not expressed. Several alleles of the pseudogene were identified in the target population. Because susceptibility to the TSEs depends on the presence of the native prion protein, an untranslated pseudogene is not expected to affect disease resistance. However, selection of genotyping methods specific for the functional gene is critical for large scale studies to identify the role of the PRNP gene in susceptibility to CWD in mule deer.