Molecular Sexing and Species Detection of Antlered European Hunting Game for Forensic Purposes

Abstract

Molecular sexing techniques are widely applied in conservation biology, although the range of forensically validated methods is fairly limited. The primary aim of this work was to develop forensically validated assays, using two PCR panels for sex and species assignment for the abundant antlered European game species: red deer (Cervus elaphus), roe deer (Capreolus capreolus) and fallow deer (Dama dama). Segments of the SRY and Amelogenin X/Y genes for sex determination, additionally species-specific cytochrome b regions for species detection were targeted and separately amplified in two multiplex reactions. These assays can reliably analyze trace amounts of DNA. The results of both can easily be visualized and interpreted practically, either on agarose gel or by capillary electrophoresis. These simple, fast molecular assays are able to affect the early-stage resolution of disputed or unsolved poaching cases, without the need of individualization or sequencing of forensic samples.

Keywords: Amelogenin X/Y; SRY; cytochrome b; fallow deer; molecular sex determination; red deer; roe deer; species detection; wildlife forensic genetics.

Figure 1

Multiplex PCR amplification for three cervid species. DeerSex-plex panel contains Amelogenin markers (194 bp for X and 140/149 bp for Y allele) and SRY (113 bp), the males show three specific bands and females show a single specific band in the agarose gel. DeerSpec-plex panel contains species-specific cytochrome b markers for roe deer (218 bp), red deer (176 bp) and fallow deer (162 bp) and show a single species specific band on agarose gel. L: ladder, m: male sample, f: female sample.

Figure 2

Parallel capillary electrophoresis detection of the PCR products amplified separately by DeerSex-plex and DeerSpec-plex assays for three cervid species. AmelY Ce: Amelogenin Y marker specific for red deer, AmelY Cc Dd: Amelogenin Y marker specific for roe deer and fallow deer, Cytb Dd: cytochrome b marker specific for fallow deer (Dama dama), Cytb Ce: cytochrome b marker specific for red deer (Cervus elaphus), Cytb Cc: cytochrome b marker specific for roe deer (Capreolus capreolus).