TaqMan Custom SNP Genotyping Assays for Forensic DNA Phenotyping of Eye Color from Saliva Stains

Single nucleotide polymorphisms (SNPs) can provide useful DNA evidence by evaluating genetic variation for phenotype prediction for criminal case investigation. The nature of SNPs allows for results to be obtained in severely degraded DNA that may not produce viable results via standard STR DNA profiling. SNP analysis by private forensic laboratories is commonly used but little published validation exists to aid one in determining the type of sample to submit or the chance of success using a SNP assay. Two key SNPs have been identified in blue/brown eye color determination: SNP rs12913832 located in the HERC2 gene and SNP rs1800407 located in the OCA2 gene. The IrisPlex software system allows for the prediction of eye color by inputting the genotypes at these SNPs obtained from custom TaqMan assays. This study focuses on eye-color determination as a model SNP system using saliva stain samples exposed to different laundering variables. Saliva samples from human donors of brown, blue, and hazel eyes were collected and used to stain fabric treated with one of the following conditions: unwashed, cold water only, cold water and bleach, hot water, and hot water and bleach. Custom Taqman assays were successful (100%) for unwashed saliva on fabric samples and partially successful for samples washed (16%) in cold water but not enough DNA was recovered for SNP testing from the other conditions. For blue and brown eyes, the IrisPlex system predicted accurately 100% of the time.