A closer look at Verogen's Forenseq™ DNA Signature Prep kit autosomal and Y‐STR data for streamlined analysis of routine reference samples

Moreno, Lilliana I.; Galusha, Michelle B.; Just, Rebecca S.

doi:10.1002/elps.201800087

Cited by 38 publications

(32 citation statements)

References 22 publications

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“…2 and S1). In general, results for the ForenSeq DNA Signature Prep Kit aSTR sensitivity were comparable to CE-based STR kits [58][59][60][61] and in line with earlier published MPS studies [1,3,4,14,54].…”

Section: Sensitivitysupporting

confidence: 88%

“…With the emergence of massively parallel sequencing (MPS) technologies, molecular genetic tools have been developed to characterise the nucleotide sequence of short tandem repeat (STR) markers that have so far been analysed using fragment sizing by capillary electrophoresis (CE) [1][2][3][4][5][6][7]. As is common practise in forensic genetics, such novel tools undergo detailed evaluation [3,[8][9][10][11][12] and validation [1,13,14] prior to their application in casework. Recent population studies have shown that sequencing of STRs lead to an increased power of discrimination compared to commonly used CE sizing by a) illustrating micro-variant structures and b) identifying sequence variations located in the repeat-(isometric variants; alleles of identical size but different in internal sequence) or flanking region [15][16][17][18][19][20].…”

Section: Introductionmentioning

confidence: 99%

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Inter-laboratory study on standardized MPS libraries: evaluation of performance, concordance, and sensitivity using mixtures and degraded DNA

et al. 2019

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We present results from an inter-laboratory massively parallel sequencing (MPS) study in the framework of the SeqForSTRs project to evaluate forensically relevant parameters, such as performance, concordance, and sensitivity, using a standardized sequencing library including reference material, mixtures, and ancient DNA samples. The standardized library was prepared using the ForenSeq DNA Signature Prep Kit (primer mix A). The library was shared between eight European laboratories located in Austria, France, Germany, The Netherlands, and Sweden to perform MPS on their particular MiSeq FGx sequencers. Despite variation in performance between sequencing runs, all laboratories obtained quality metrics that fell within the manufacturer's recommended ranges. Furthermore, differences in locus coverage did not inevitably adversely affect heterozygous balance. Inter-laboratory concordance showed 100% concordant genotypes for the included autosomal and Y-STRs, and still, X-STR concordance exceeded 83%. The exclusive reasons for X-STR discordances were drop-outs at DXS10103. Sensitivity experiments demonstrated that correct allele calling varied between sequencing instruments in particular for lower DNA amounts (≤ 125 pg). The analysis of compromised DNA samples showed the drop-out of one sample (FA10013B01A) while for the remaining three degraded DNA samples MPS was able to successfully type ≥ 87% of all aSTRs, ≥ 78% of all Y-STRs, ≥ 68% of all X-STRs, and ≥ 92% of all iSNPs demonstrating that MPS is a promising tool for human identity testing, which in return, has to undergo rigorous in-house validation before it can be implemented into forensic routine casework.

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Section: Sensitivitysupporting

confidence: 88%

Section: Introductionmentioning

confidence: 99%

Inter-laboratory study on standardized MPS libraries: evaluation of performance, concordance, and sensitivity using mixtures and degraded DNA

et al. 2019

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“…sequencing error, PCR error, DNA damage, pseudogenes) could be consistently distinguished from authentic signal. We have employed methods for doing exactly that in other NGS studies [23,56,57]. However, these analyses have been largely manual, which is not suitable to a production environment.…”

Section: Discussionmentioning

confidence: 99%

Validation of NGS for mitochondrial DNA casework at the FBI Laboratory

Brandhagen

Just

Irwin

2020

Forensic Science International: Genetics

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As a first step towards integrating next generation sequencing (NGS) technology into the FBI Laboratory's operational casework, the PowerSeq™ CRM Nested System, an NGS-based mitochondrial DNA (mtDNA) control region assay, was developmentally and internally validated. The validation studies were conducted in accordance with the Scientific Working Group on DNA Analysis Methods (SWGDAM) Validation Guidelines for Forensic DNA Analysis Methods, and the FBI's Quality Assurance Standards (QAS) for Forensic DNA Testing Laboratories. The assay was shown to be highly reproducible, with variant frequencies across intra and inter-run replicates of the same sample differing, on average, by just 0.3% for substitutions and point heteroplasmies and 1.5% for insertions and deletions. The assay was also shown to be extremely sensitive, yielding complete control region sequence data from as few as 2000 copies of mtDNA. This is a more than 20-fold increase in sensitivity when compared to the FBI Laboratory's current Sanger sequencing-based protocols and, based on mtDNA quantitation values of samples routinely encountered in mtDNA casework, suggests that the percentage of questioned samples from which full control region data can be recovered will increase from our current 20% to approximately 90% success with NGS technology. In addition, the assay requires on average only 30% of the extract volume typically required to develop control region profiles from degraded samples via Sanger sequencing. Overall, these studies establish the reliability of the PowerSeq™ CRM Nested System for accurate mtDNA control region typing and can serve as a model for laboratories seeking to validate NGS protocols for forensic mtDNA analysis.

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“…The large-scale performances and functional tests had been widely validated for the ForenSeq™ DNA Signature Prep Kit, the DNA Primer Set A (DPMA, 152 loci totally: 27 common, forensic autosomal STRs, 24 Y-STRs, 7 X-STRs, and 94 identity informative SNPs) and the DNA Primer Set B (in total 230 loci: DMPA plus 22 phenotypic informative SNPs and 56 biogeographical ancestry SNPs), which also include robustness, reproducibility, species specificity, consistency and sensitivity of detection [40][41][42][43][44][45][46][47]. In addition, the applicability of the DNA Signature Prep Kit had been evaluated and verified on challenging samples, DNA mixture, degradative DNA [48][49][50], formalin-fixed paraffin-embedded tissues [49,51], and remains of skeletons and bones [48,[52][53][54], to extend the applications for forensic sciences [55][56][57][58][59][60][61][62][63][64][65][66][67][68]. Hence, on the basis of the sufficient validations of the stability and homogeneity, more essential data of worldwide populations are needed urgently at present to improve the standard experimental procedures, promote the efficiencies of MPS-based system, and strengthen and unify the contacts between capillary electrophoresis-based (CE-based) and MPS-based data for the basic and extended applications of forensic sciences and others.As the aborigines in Hainan island where was the entrances to East Asia (either the southern entrance from Indo-China peninsula or the northern entrance from Central Asia) [12,[69][70][71], Hainan...…”

mentioning

confidence: 99%

The forensic landscape and the population genetic analyses of Hainan Li based on massively parallel sequencing DNA profiling

Fan

Wang

et al. 2020

Preprint

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Due to the formation of the Qiongzhou Strait by climate change and marine transition, Hainan island isolated from the mainland southern China during the Last Glacial Maximum. Hainan island, located at the southernmost part of China and separated from the Leizhou Peninsula by the Qiongzhou Strait, laid on one of the modern human northward migration routes from Southeast Asia to East Asia. The Hlai-language speaking Li minority, the second largest population after Han Chinese in Hainan island, is the direct descendants of the initial migrants in Hainan island and has unique ethnic properties and derived characteristics, however, the forensic associated studies on Hainan Li population are still insufficient.Hence, 136 Hainan Li individuals were genotyped in this study using the MPS-based ForenSeq™ DNA Signature Prep Kit (DNA Primer Set A) to characterize the forensic genetic polymorphism landscape, and DNA profiles were obtained from 152 different molecular genetic markers (27 autosomal STRs, 24 Y-STRs, 7 X-STRs, and 94 iiSNPs). A total of 419 distinct length variants and 586 repeat sequence sub-variants, with 31 novel alleles (at 17 loci), were identified across the 58 STR loci from the DNA profiles of Hainan Li population. We evaluated the forensic characteristics and efficiencies of DAPA, it demonstrated that the STRs and iiSNPs in DAPA were highly polymorphic in Hainan Li population and could be employed in forensic applications. In addition, we set up three Datasets, which included the genetic data of (Ⅰ). iiSNPs (27 populations, 2640 individuals), (Ⅱ). Y-STRs (42 populations, 8281 individuals), and (Ⅲ). Yhaplogroups (123 populations, 4837 individuals) along with the population ancestries and language families, to perform population genetic analyses separately from different perspectives.In conclusion, the phylogenetic analyses indicated that Hainan Li, with a southern East Asia origin and Tai-Kadai language-speaking language, is an isolated population relatively. But the genetic pool of Hainan Li influenced by the limited gene flows from other Tai-Kadai populations and Hainan populations. Furthermore, the establishment of isolated population models will be beneficial to clarify the exquisite population structures and develop specific genetic markers for subpopulations in forensic genetic fields. Signature Prep Kit; Massively parallel sequencing. mass landings of the Han (since BC 214 in Qin Dynasty) and Lingao people (about BC 500 during the Spring and Autumn period) compressed the living space of Hainan Li, making Li from the north to the south of Hainan island [1,2,9,29,30]. (Nowadays, the main distributions of Hainan Li minority are shown in Fig. 1C.) In all ages, ever since the primordial settlements in Hainan island, Hlai language was the dominant language for the original migrants and their descendants, and the intangible cultural and spiritual heritages within Hainan Li were preserved by oral transmission without ancient writings to hand down. Hlai, which is one important branch of the Tai-Kadai languag...

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A closer look at Verogen's Forenseq™ DNA Signature Prep kit autosomal and Y‐STR data for streamlined analysis of routine reference samples

Cited by 38 publications

References 22 publications

Inter-laboratory study on standardized MPS libraries: evaluation of performance, concordance, and sensitivity using mixtures and degraded DNA

Inter-laboratory study on standardized MPS libraries: evaluation of performance, concordance, and sensitivity using mixtures and degraded DNA

Validation of NGS for mitochondrial DNA casework at the FBI Laboratory

The forensic landscape and the population genetic analyses of Hainan Li based on massively parallel sequencing DNA profiling

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