Gender determination of forensic samples using PCR amplification of ZFX/ZFY gene sequences

Determining the gender of an evidentiary sample can be an important part of casework analyses. Gender information, particularly when combined with mitochondrial DNA analysis, can serve to distinguish biological evidence from two people who share the same DNA type(s) but differ by sex. When typing sexual assault evidence, gender information can serve as confirmation that the "sperm fraction" extracted from swabs and stains actually contains male DNA and also as an indicator of the amount of male DNA present in the non-sperm fraction. The PCR-based assay described here relies on amplification of a small, polymorphic region of a homologous zinc finger protein locus present on the X and Y chromosomes. The gender of the sample donor is determined from the PCR product either by Haelll restriction enzyme digestion followed by gel electrophoresis or by hybridization to immobilized sequence specific oligonucleotide probes (reverse dot blot). When using the reverse dot blot approach, amplification and typing of the gender PCR product can be coupled to amplification and typing of the AmpliType HLA DQalpha and PM markers. Sensitivity and mixture studies were performed in addition to the analysis of casework bloodstains and sexual assault kit samples. Additional studies using this gender determination assay are described in the accompanying paper.