The SeqStudio™ for human identification (HID) is a new benchtop capillary electrophoresis (CE) platform recently developed by Applied Biosystems for genotyping and sequencing short tandem repeat (STR) fragments. Compared to the previous series of CE systems developed by this maker, it is more compact and easier to use. Moreover, by allowing the detection of 4 to 8 fluorescent dyes, it seems to be fully compatible with the different kits of autosomal and gonosomal STR markers usually used in forensic genetics, which are available in trade and supplied by various manufacturers. However, being a new CE model, before its routine use in forensic genetics applications, it should undergo appropriate analytical validation studies in its own laboratories to understand its potential and limitations. A series of experiments on DNA samples coming from cell line controls, using the GlobalFiler™ IQC Amplification Kit, were carried out to meet this purpose. The SeqStudio™ Genetic Analyzer for HID\'s findings on genotyping reproducibility (precision and accuracy of sizing), sensitivity, signal variability between dyes (intra- and inter-color channel balance), and stutter ratios are reported. These findings confirm the validity of this new CE system and its capability to generate reliable results.

The STRtyper-32G PCR Amplification Kit is a 6-dye multiplex system that combines the 30 autosomal STR loci with an Indel site (YIndel) and the sex-determinant locus Amelogenin. In addition to more loci, Master Mix has been optimized to amplify DNA on different substrates. The autosomal STR loci contained in this novel system meet the compatibility of requirements for databasing. In this study, the developmental validation study of the STRtyper-32G Kit followed the guidelines of SWGDAM (Scientific Working Group on DNA Analysis Methods), including PCR-based studies, species specificity, inhibitors, sensitivity, precision, repeatability, stutter, DNA mixtures, concordance studies, and population genetics studies. The validation results indicate that the new multiplex system is a robust tool for forensic database applications.

The VeriFiler™ Plus PCR Amplification Kit is a 6-dye multiplex assay that simultaneously amplifies a set of 23 autosomal markers (D3S1358, vWA, D16S539, CSF1PO, D6S1043, D8S1179, D21S11, D18S51, D5S818, D2S441, D19S433, FGA, D10S1248, D22S1045, D1S1656, D13S317, D7S820, Penta E, Penta D, TH01, D12S391, D2S1338, and TPOX), a quality indicator system, and two sex-identification markers. Combined, the markers satisfy the requirements of the Chinese National autosomal DNA database as well as expanded CODIS (Combined DNA Index System). The VeriFiler Plus kit was developed with an improved Master Mix which incorporates the brighter TED™ dye, and accommodates a higher sample loading volume thus allowing for increased sensitivity and enabling maximum information recovery from challenging casework samples including touch, degraded, and inhibited samples. Here, we report the results of the developmental validation study which followed the SWGDAM (Scientific Working Group on DNA Analysis Methods) guidelines and includes data for PCR-based studies, sensitivity, species specificity, stability, precision, reproducibility and repeatability, concordance, stutter, DNA mixtures, and performance on mock casework samples. The results validate the multiplex design as well as demonstrate the kit\'s robustness, reliability, and suitability as an assay for human identification with casework DNA samples.

The Huaxia™ Platinum Kit for short tandem repeat (STR) amplification was designed to meet the needs of the rapidly growing Chinese forensic database. This PCR multiplex allows simultaneous amplification of the following autosomal loci: D3S1358, vWA, D16S539, CSF1PO, TPOX, D8S1179, D21S11, D18S51, Penta E, D2S441, D19S433, TH01, FGA, D22S1045, D5S818, D13S317, D7S820, D6S1043, D10S1248, D1S1656, D12S391, D2S1338, Penta D and the gender-identification markers Yindel, and AMEL. The Huaxia™ Platinum Kit enables direct amplification from blood and buccal samples stored on treated and untreated paper, and features an optimized PCR protocol that yields time to results in less than 45 min. Developmental validation testing followed SWGDAM guidelines and demonstrated that this assay produces reproducible and accurate results. Studies on 798 individuals in 4 major Chinese ethnic groups produced highly concordant results with other commercially available STR genotyping kits. The validation results demonstrate that the Huaxia™ Platinum Kit is a robust and reliable identification system for forensic DNA databasing applications.

The GlobalFiler™ PCR Amplification Kit is a single multiplex assay that amplifies a set of 24 markers, which encompass the European Standard Set and CODIS (Combined DNA Index System) recommended composite set of loci. In addition to more loci and a 6-dye chemistry format, the Master Mix has been formulated to allow higher sample loading volume for trace DNA samples. The GlobalFiler™ Kit has been optimized to deliver high performance on casework samples, while also delivering fast thermal cycling, with an amplification time of approximately 80 min. Here, we report the results of the developmental validation study which followed the SWGDAM (Scientific Working Group on DNA Analysis Methods) guidelines and includes data for PCR-based studies, sensitivity, species specificity, stability, precision, reproducibility and repeatability, concordance, stutter, DNA mixtures, and performance on mock casework samples. The results validate the multiplex design as well as demonstrate the kit\'s robustness, reliability, and suitability as an assay for human identification with casework DNA samples.

The original CODIS database based on 13 core STR loci has been overwhelmingly successful for matching suspects with evidence. In order to increase the power of discrimination, reduce the possibility of adventitious matches, and expand global data sharing, the CODIS Core Loci Working Group determined the expansion of the CODIS core loci to 20 STR plus three additional \"highly recommended\" loci (SE33, DY391, Amelogenin) Hares, 2015, 2012 [1,2]. The QIAGEN Investigator 24plex QS and Investigator 24plex GO! Kits are 6-dye multiplex assays that contain all markers of the expanded 23 CODIS core loci along with a unique internal performance control that is co-amplified with the STR markers. The \"Quality Sensor\" generates additional information for quality control and performance checks. Investigator 24plex QS is designed for purified DNA from casework and reference samples, whereas 24plex GO! is dedicated to direct amplification of reference samples, like blood or buccal cells on FTA or swabs. A developmental validation study was performed on both assays. Here, we report the results of this study which followed the recommendations of the European Network of Forensic Science Institutes (ENFSI) [3] and the Revised Validation Guidelines of the Scientific Working Group on DNA Analysis Methods (SWGDAM) [4]. Data included are for PCR-based procedures e.g. reaction conditions, effects of PCR annealing temperature variations, amplification cycles or cyclers, sensitivity (also in the context of the Quality Sensor), performance with simulated inhibition, stability and efficiency, precision, reproducibility, mixture study, concordance, stutter, species specificity, and case-type samples. The validation results demonstrate that the Investigator 24plex QS and Investigator 24plex GO! Kits are robust and reliable identification assays as required for forensic DNA typing in forensic casework analysis and databasing.

In order to increase the power of discrimination, reduce the possibility of adventitious matches, and expand global data sharing, the CODIS Core Loci Working Group made a recommendation to expand the CODIS core loci from the \"required\" 13 loci to 20 plus three additional \"highly recommended\" loci. The GlobalFiler(®) Express Kit was designed to incorporate all 20 required and 3 highly recommended loci along with a novel male-specific Y insertion/deletion marker. The GlobalFiler(®) Express Kit allows simultaneous amplification of the following loci: D3S1358, vWA, D16S539, CSF1PO, TPOX, Yindel, AMEL, D8S1179, D21S11, D18S51, DYS391, D2S441, D19S433, TH01, FGA, D22S1045, D5S818, D13S317, D7S820, SE33, D10S1248, D1S1656, D12S391, and D2S1338. The kit enables direct amplification from blood and buccal samples stored on paper or swab and the chemistry features an optimized PCR protocol that yields time to results in less than an hour. Developmental validation testing followed SWGDAM guidelines and demonstrated the quality and robustness of the GlobalFiler(®) Express Kit over a number of variables. The validation results demonstrate that the 24-locus multiplex kit is a robust and reliable identification assay as required for forensic DNA typing and databasing.

The idea of developing a new multiplex STR amplification system was conceived in 2011 as an effective way to implement the new European standard set (ESS) of 12 STR markers adopted by The Council of the European Union in 2009 while maintaining an effective compatibility and information exchange with the historical DNA profiles contained in the Spanish national DNA database (around 200,000 DNA profiles) mainly based on the 13 CODIS core STR loci plus D19S433 and D2S1338 markers. With this goal in mind we proposed to test and validate a single STR amplification system for simultaneous analysis of 21 STR markers covering both CODIS and ESS core STR loci plus three additional markers (D19S433, D2S1338, and SE33) also contained in commonly used STR kits and national DNA databases. In 2012, we started the first beta-testing with a 6-dye STR kit prototype containing 24 loci (now known as the GlobalFiler™ PCR Amplification Kit) developed by Life Technologies in response to the CODIS Core Loci Working Group\'s recommendation to expand the CODIS Core Loci. This prototype included our proposal of 21 autosomal STR markers and two Y-chromosome markers (DYS391 and Y-indel) and maximizes concordance with established databases and previously analyzed samples by maintaining primer sequences of previous Identifiler(®)/NGM SElect™ kits for the 21 STR markers except for TPOX. This paper describes the validation studies conducted with the first commercial available 6-dye STR kit for casework using a 3500 genetic analyzer for fragment detection that included the analysis of the following parameters and aspects: analytical threshold, sensitivity & stochastic threshold, heterozygous balance, stutter threshold, precision and accuracy, repeatability and reproducibility, genotype concordance, DNA mixtures, species specificity, and stability studies with case type samples. The studies demonstrated that the GlobalFiler™ system provided equivalent overall performance to previous forensic STR PCR kits, but with enhanced discrimination power for a better match efficiency that would reduce the chance of adventitious matches during DNA data exchange among national DNA databases.