Comparison of EMA Decontamination of Different Primer Sets.

<p>Three primer sets of increasing amplicon length were treated with EMA and assessed for positivity in no template controls and detection of low level spiked <i>E</i>. <i>coli</i> gDNA. 24 no template controls (NTC) and 8 positive reactions of 20 genome copies (20c) and 2 genome copies (2c) each were performed for each EMA concentration per primer set.</p><p>Comparison of EMA Decontamination of Different Primer Sets.</p

Sequences of Primers Used in this Study.

<p>Nucleotide positions correspond to <i>E</i>. <i>coli</i> sequence (Genbank accession number J01859). Annealing temperatures and extension times are given for each primer pair.</p><p>Sequences of Primers Used in this Study.</p

UV Decontamination of SF3c-SR5 Primer Set.

<p>Master mixes of all reaction components (excluding EvaGreen dye) were exposed to UV irradiation before the addition of dye and template (PCR water in no template controls, and 20 or 2 <i>E</i>. <i>coli</i> genome copies for positives). 42 no template control reactions and 32 positive reactions (16 per template amount) were performed for each UV condition, with 22 NTC and 16 positive reactions respectively for non-treated controls. Reactions that did not amplify within the 40 cycle threshold are represented as 40 for visual comparison. Effects of treatments on the number of positive NTC reactions, as compared to no treatment controls, were analysed statistically by Fisher’s exact test. * = P <0.0001. Horizontal bars = median value; percentages = number of positive reactions.</p

Separate EMA Treatment of Primers.

<p>Reactions were repeated with primers added before a two minute UV exposure, compared to post-UV addition of primers treated with or without 1μM EMA. 36 NTC reactions and 12 positive reactions per template concentration were performed for each method. Reactions that did not amplify within the 40 cycle threshold are represented as 40 for visual comparison. Percentages are given for the number of reactions giving a positive result within the 40 cycle threshold. Effects of primer treatments on the number of positive NTC reactions, as compared to non-primer treated controls, were analysed statistically by Fisher’s exact test. * = P <0.0001.</p

Detectable levels of PrP^Sc on Western blots do not correlate with the levels of infectivity.

<p>10% brain homogenates from an uninfected brain (lane 2) time-course samples week 3, 6, 9, 12, 15, 18, 21 (lane 3–9), and the terminal sample (lane 10) were digested with proteinase K at 60°C for 10 minutes and assessed by Western blot. The observed signal does not correspond with the levels of infectivity found in corresponding bioassays for the week 12–21 post-exposure time-points.</p

Comparison of the cull dates for the mice challenged via the gingival margin.

<p>Panel A; Frequency distribution plots show the presence of a normally distributed population with a mean incubation period of around 250 days plus a small number of animals with significantly shorter incubations ranging from 140–188 days. Panel B; when these two groups are compared they show distinct means and distribution and are considered as distinct populations (p<0.001).</p