Theory and practice of prion protein analysis in food products
2014
Prosekov, A.Yu., Kemerovo Inst. of Food Science and Technology (Russian Federation)
The results of the research on methods of identification and quantitative determination of prion proteins in biological samples and multicomponent mixtures based on them are presented. Analysis of nucleotide sequence of DNA encoding the PRNP gene of the prion protein, including phylogenetic and comparative analysis of nucleotide sequences of normal and pathogenic prion protein in cattle, was performed. Oligonucleotide primers for amplification of the PRNP gene of pathogenic prion protein were designed and synthesized. The higher stability of detection of low DNA concentrations in the PCR method, if compared with other methods, is confirmed by the results of clinical samples study. Comparison of the results obtained with the proposed PCR test-system and the reference method evidenced real-time high specificity of the developed PCR method. The high specificity of the developed test system was confirmed by three ways: 1) using the Primer3 software; 2) by electrophoretic separation of the meat chop samples with different percent content of pork tissues infected with a pathogenic prion protein; and 3) by comparative analysis of the results of pathogenic prion protein determination using the proposed PCR test-system and a commercial ELISA assay TeSeE.
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