Analysis of nucleic acids in purified scrapie prion preparations

Summary

Amount, type, and size of nucleic acid molecules associated with purified prion preparations were analyzed. Return refocusing gel electrophoresis (RRGE) was developed to detect homogeneous and heterogeneous nucleic acids extracted from highly purified scrapie prion preparations. With this method all types of nucleic acids in the size range from 13 to several thousand nucleotides could be analyzed. The recovery of all nucleic acids, after deproteinization and two-phase extraction was higher than 90%. Despite extensive nuclease digestions some small polynucleotides remained. Although a scrapie-specific nucleic acid cannot be excluded, the results further define the possible characteristics for such a hypothetical molecule. If it was homogeneous in size, then it would be <80 nt in length at a particle-to-infectivity ratio (P/I) near unity; if the other extreme, i.e. totally heterogeneous scrapiespecific nucleic acids were assumed, then scrapie-specific nucleic acids would have to include molecules smaller than 240 nt. In order to exclude the possibility that unspecific background nucleic acid is entrapped in prion-rods, infectious material has to be prepared without a proteolysis and rod formation, and the analysis of nucleic acids performed with those preparations.