Characterization and validation studies of powerPlex 2.1, a nine-locus short tandem repeat (STR) multiplex system and penta D monoplex

In order to increase the power of discrimination for human identification purposes, a nine-locus short tandem repeat (STR) multiplex, the GenePrint PowerPlex 2.1 system (PowerPlex 2.1) developed by Promega Corporation and a separate pentanucleotide-repeat locus, Penta D, were tested. This megaplex system includes the highly polymorphic loci FGA, TPOX, D8S1179, vWA, Penta E, D18S51, D21S11, TH01, and D3S1358 and may be used in combination with the eight-locus STR multiplex, the GenePrint PowerPlex 1.1 system (PowerPlex 1.1) that has been previously developed. Three of the loci, TPOX, TH01 and vWA, have been included in both systems for quality control purposes. As with PowerPlex 1.1, PowerPlex 2.1 is also based on a two-color detection of fluorescent-labeled DNA products amplified by polymerase chain reaction (PCR) and provides a valuable tool for accurate and rapid allele determination. The primer sequences used in the PowerPlex 2.1/Penta D system are also presented in this report. To meet the "Quality Assurance Standards for Forensic DNA Testing Laboratories" (FBI), we tested the efficiency and reproducibility of the PowerPlex 2.1/PentaD system by several validation studies that were conducted as a joint project among seven laboratories. Validation tests included concordance studies, sensitivity, and species specificity determination, as well as performance in forensic and environmentally impacted samples. The results produced from these tests demonstrated the consistency and reliability of the PowerPlex 2.1/Penta D system.     

Validation and implementation of the PowerPlex 16 BIO System STR multiplex for forensic casework

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 DS5S818), 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.