Discrimination Between Human and Animal DNA: Application of a Duplex Polymerase Chain Reaction to Forensic Identification

Identification of a report's species is one of the basic analyses in forensic laboratories. The authors report the case of 6 bone fragments recovered in a wooded area, which were not attributable to 1 animal species on the basis of morphologic examination. The aim of this study was to develop a duplex polymerase chain reaction (PCR) to discriminate human and animal origin of bone fragments. The method is based on the PCR amplification of cytochrome b and a 16S ribosomal mitochondrial DNA fragment, which has never been tested up to now. Our protocol combines a single-round PCR with direct visualization of amplicons in agarose gel, without sequencing analysis of the PCR products. The presence of a single band (359 bp) indicates a nonhuman origin of the sample, whereas 2 bands (157 and 359 bp) indicate a human biologic sample.This method revealed to be useful for forensic purposes because the 16S ribosomal mitochondrial DNA is a small human-specific fragment that is easily amplifiable even with degraded DNA from biologic materials such as old bones.