Detection of nucleic acids to assess risk for bovine spongiform encephalopathy
View Patent ↗The present invention provides a method of detecting abnormal serum nucleic acid profiles to assess the risk of a transmissible spongiform encephalopathy, e.g., BSE.
1. A method of detecting an animal at increased risk for bovine spongiform encephalopathy (BSE), the method comprising:
incubating nucleic acids extracted from an acellular sample obtained from the animal with amplification primers in a test amplification reaction; wherein the amplification primers comprise: (i) at least 10 contiguous nucleotides of CHX-1F (SEQ ID NO:1) and CHX-1R (SEQ ID NO:2) or (ii) CHX-1F (SEQ ID NO:1) and CHX-1R (SEQ ID NO:2) or (iii) 63R (SEQ ID NO: 11) and 55F (SEQ ID NO: 3); or 60F (SEQ ID NO: 5) and 55R (SEQ ID NO: 4);
detecting reactivity of the amplification reaction that is over 5 standard deviations from a reference amplification reaction, wherein reactivity of over 5 is indicative of an increased risk for BSE, wherein reactivity of the amplification reaction is determined using a melting curve.
2. The method of claim 1 , wherein the acellular fluid sample is serum.
3. The method of claim 1 , wherein the acellular fluid sample is plasma.
4. The method of claim 1 , wherein one of the primers comprises sequences that hybridize to repetitive sequences in the animal genome.
5. The method of claim 4 , wherein the repetitive sequences are SINE sequences.
6. The method of claim 1 , wherein the nucleic acid sample comprises DNA.
7. The method of claim 1 , wherein the amplification reactions comprise a compound that specifically binds to double-stranded DNA.
8. The method of claim 7 , wherein the compound is a fluorescent dye.
9. The method of claim 1 , wherein the amplification primers comprise CHX-1F (SEQ ID NO:1) and CHX-1R (SEQ ID NO:2).
10. The method of claim 1 , wherein the amplification primers comprise 63R and 55F; or 60F and 55R.