In this work four species-specific primers and probes were designed and evaluated for the detection and quantification of bovine, ovine, swine
and chicken mitochondrial DNA in feeds. PCR primers were optimized using conventional and Real Time PCR, to detect short species-specific
sequences amplifiable from heat treated material. Both methods confirmed the high specificity of the primers designed. Real time quantitative
PCR assay allowed the detection of as few as 0.01 ng and 0.05 ng of ovine and bovine genomic DNA, respectively. The detection limit for swine
and chicken genomic DNA was 0.5 ng. Sensitivity levels observed in DNA extracted from meat samples processed according to EU legislation
were different compared to those in genomic DNAs previously described. They resulted in swine 5 fg of MBM DNA, in chicken 25 ng, in ovine
and bovine 50 ng. We confirmed the efficiency and specificity of primers in RT-PCR to detect 0.5% of bovine, ovine, swine and chicken MBM in
contaminated feedstuffs.
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