Developmental validation of the Microreader™ RM-Y ID System: a new rapidly mutating Y-STR 17-plex system for forensic application.

Y-chromosomal short tandem repeats (Y-STRs) are widely applied to evolutionary, genealogical, and kinship analyses of male linages in forensic studies, but these low to midrange mutated Y-STRs typically fail to separate related males from the same paternal lineage. Recently, rapidly mutating Y-STRs (RM Y-STRs) have been demonstrated to improve the differentiation of male relatives and individuals. The Microreader™ RM-Y ID System is a new RM Y-STR kit that is capable of simultaneously amplifying 17 RM Y-STRs. Herein, to verify the efficiency and accuracy of the Microreader™ RM-Y ID System, developmental validation was conducted, including PCR-based studies, sensitivity, stability, species specificity, mixture, stutter percentage, and precision studies. Full profiles could be obtained when the hematin concentration was 250 μM, humic acid concentration was 1500 ng/μl, and tannic acid concentration was 200 ng/μl. Full profiles of the mixture of males/males could be detected up to a ratio of 19:1, and full profiles of females/males could always be detected even at ratios up to 24,000:1. Moreover, the forensic characteristics of 250 DNA-confirmed father-son pairs were analysed. The results showed that these 17 RM Y-STRs had high power for forensic discrimination (HD = 1) in the Chinese Han population, and the mutation rates were in the range of 4 × 10 (95% CI 1.00 × 10 to 2.21 × 10, DYS464) to 8.8 × 10 (95% CI 5.60 × 10 to 1.30 × 10, DYF399S1), indicating that the kit was effective for RM Y-STR studies and absolute individualisation of interrelated male individuals.