This document specifies a real-time PCR procedure for the quantitation of the amount of equine DNA relative to total mammalian DNA in a raw meat sample.
Results of this equine assay are expressed in terms of equine (Equus genus) haploid genome copy numbers relative to total mammalian haploid genome copy numbers. This assay is specific for representatives of the genus Equus and therefore detects horse, mule, donkey and zebra DNA.
The method has been previously validated in a collaborative trial and applied to DNA extracted from samples that consist of raw horse meat in a raw beef (meat) background.
The limit of detection has been determined experimentally to be at least 9 genomic equivalent copies (~ 17 haploid target gene copies) for both the horse genome (E. caballus) and mammalian genome (raw meat samples) based on the lowest dilution on the respective calibration curves through single laboratory validation. The lowest relative horse content of the target sequence included in the collaborative trial was a mass fraction of 0,1 % based on gravimetrically prepared raw horse muscle tissue in a raw beef muscle tissue background.
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