Study of male DNA quantification methods in male-female mixture samples

Abstract

 DNA quantification is an important process in DNA typing. Today, quantitative real-time PCR (qPCR) targeting the autosomal D17Z1 region, which cannot distinguish between male and female DNA, is the main method for forensic human DNA quantification for the Japanese police. For more efficient testing Y-STR type, we examined the forensic evaluation (sensitivity, repeatability, specificity, inhibition, comparison with commercial qPCR reagent and simulated mixture study) of qPCR assays targeting Y-chromosomal Sex determining region Y (SRY) and DYZ5 regions. We evaluated the effectiveness of simple and practical male DNA quantification methods in male and female mixture samples. The sensitivity of SRY and DYZ5 assays were 0.015 ng/μL and 0.0015 ng/μL. Both assays could quantify male DNA in the presence of high concentrations of female DNA. The SRY assay had the same sensitivity as the commercial qPCR reagent, but had more resistant to inhibitors. The DYZ5 assay tended to have higher DNA concentration than the commercial qPCR reagent and the SRY assay, but could detect trace amounts of male DNA. The reason for this is that DYZ5 region is multi-copy locus and there are individual differences in the copy number. The DYZ5 assay was considered to be one of the effective methods for proving the presence of male DNA in the sample. Therefore, we consider that these assays can select the regions according to purpose and utilized to forensic applications.