Validation and Implementation of the PowerPlex® 16 BIO System STR Multiplex for Forensic Casework

Abstract

The PowerPlex® 16 BIO multiplex short tandem repeat (STR) system contains the 13 CODIS loci (FGA, TPOX, D8S1179, vWA, D18S51, D21S11, TH01, D3S1358, CSF1PO, D16S539, D7S820, D13S317, and D5S818), plus two pentanucleotide repeat loci (Penta D and Penta E) and the sex-identifying locus, Amelogenin. The PowerPlex® 16 BIO System is optimized for use with the Hitachi FMBIO® gel imaging systems. A consortium of seven independent laboratories collaborated to perform the studies defined by the FBI standards for performing a developmental validation, including the evaluation of sample concordance, percent stutter determination, nonprobative casework, precision, sensitivity, mixture determination, effect of substrates, the impact of environmental insults, and species specificity. All samples tested for concordance were consistent except for one sample from the Virginia Division of Forensic Science database that displayed discordance at D13S317, a locus whose primer sequence was altered. Stutter values were comparable to those of other STR multiplex systems, the precision was comparable to other multiplexes analyzed by gel electrophoresis, the DNA profiles were unchanged by the substrate upon which the blood samples were placed, and the nonprobative casework samples re-typed for the PowerPlex® 16 BIO System were consistent with previous typing results. When greater than 0.125 ng of DNA was placed into the PowerPlex® 16 BIO System amplification reaction, a full profile was generated by all laboratories. The mixture study results were comparable to those reported for other multiplex systems, the environmental study demonstrated a loss of larger molecular weight loci when samples were incubated at elevated temperatures for a prolonged period of time, and the only notable cross species hybridization was observed with primate DNA samples. This extensive validation work performed demonstrates that the PowerPlex® 16 BIO System provides STR data of a quality comparable with other PowerPlex® STR multiplex kits as well as other widely used STR multiplexes and is thus suitable for evidentiary casework analysis as well as database sample profiling.

A Forensic Laboratory Tests the Berkeley Microfabricated Capillary Array Electrophoresis Device*

“…(36) reported full detection of the minor contributor at the 3:1 and 1:3 ratios; however, the threshold setting was lower than that routinely applied to casework samples (50 rfu) and minor contributor alleles at the stutter position were excluded from analysis, which was not the case in the study reported here. Minor contributor alleles were above the peak threshold at many loci for both the 9:1 and 1:9 ratios as is also consistent with other reports (13,33,35,36). The dynamic coating polymer, pDuramide, was utilized for microchip coating in this experiment, which again indicated that this alternate coating procedure did not adversely impact μCAE device performance.…”

“…The majority of minor contributor alleles were observed at the 3:1 and 1:3 ratios. However, at the 3:1 ratio, a 12 allele at D13S317 was below the threshold for reporting as were a 9 allele at TH01 and an 11 allele at Penta E. These findings are not unexpected and are consistent with previously reported mixture results using commercial detection platforms (33,35). All minor contributor alleles were reported in the Yeung et al.…”

Haplotyping of STR cluster DXS6801–DXS6809–DXS6789 on Xq21 provides a powerful tool for kinship testing

“…1). This case was initially analysed using 19 autosomal markers: D3S1358, vWA, D16S539, D2S1338, D8S1179, ACTBP2, D19S433, TH01, FIBRA, D21S11, D18S51, Penta E, D5S818, D13S317, D7S820, D16S539, CSF1PO, Penta D, and TPOX [1,9]. At the same time, we initiated X-chromosomal testing and investigated the Xq21 cluster described above together with 15 additional X-chromosomal markers reported earlier [2-4, 6, 7, 10, 12, 16].…”

Fluorescence energy transfer‐labeled primers for high‐performance forensic DNA profiling

“…This multiplex was chosen for modification because of its readily available sequence information which is critical for primer design [19] and it has previously been validated for use on forensic casework evidence [20]. The ET 16-plex typing system was constructed first using the same primer concentrations in PowerPlex 16 (personal communication, Promega) followed by subsequent adjustments of primer concentrations by iteratively typing a single-source DNA sample, A-18.…”