Systematic evaluation of the Precision ID GlobalFiler™ NGS STR panel v2 using single-source samples of various quantity and quality and mixed DNA samples
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引用次数: 0
Abstract
Massively parallel sequencing (MPS) techniques were developed approximately 15 years ago. Meanwhile, several MPS kits for forensic identification, phenotypic information, ancestry, and mitochondrial DNA analysis have been developed and their use has been established. Sequencing short tandem repeats (STRs) has certain advantages over the currently used length-based genotyping methods, which are based on PCR amplification followed by capillary electrophoresis (CE). MPS is more discriminative and includes the possibility of testing high numbers of targets (> 100), different types of markers [STRs and single nucleotide polymorphisms (SNPs)], as well as the use of smaller amplicons (< 300 bp). This study evaluated in 24 experimental runs the Precision ID GlobalFiler™ NGS STR panel v2 from ThermoFisher, which targets 31 autosomal STRs, amelogenin, and three Y-markers (one STR, SRY, and Yindel). Single-source samples were used in 18 experimental runs, for systematic evaluation. These included assessing library preparation benchmark conditions, limited DNA input, as well as testing repeatability, number of samples per run, and degraded DNA samples. Full profiles were consistently obtained from as little as 50 pg DNA input. Using the optional recovery PCR method improved outcomes for samples with low DNA input. Full profiles were also obtained from severely degraded DNA samples with degradation indices (DI) of > 60. In addition, six experimental runs were performed testing various two-person mixtures with mixture ratios ranging from 1:20 to 20:1. Major and minor contributors were distinguishable by their read counts (coverage), because less DNA input yielded lower read counts, analogous to the traditional CE technology, where less DNA produces lower peak heights. Mixture ratios of approximately 1:1 were indistinguishable, while a greater imbalance, i.e., higher mixture ratios, made the mixture more distinguishable between major and minor contributors. Based on this information, the highest success rate of correctly deconvoluted four-allelic loci was from mixtures with 1:3 ratios. At higher mixture ratios, the drop-out rate of the minor contributor increased, reducing the number of four-allelic loci.
期刊介绍:
Forensic Science International: Genetics is the premier journal in the field of Forensic Genetics. This branch of Forensic Science can be defined as the application of genetics to human and non-human material (in the sense of a science with the purpose of studying inherited characteristics for the analysis of inter- and intra-specific variations in populations) for the resolution of legal conflicts.
The scope of the journal includes:
Forensic applications of human polymorphism.
Testing of paternity and other family relationships, immigration cases, typing of biological stains and tissues from criminal casework, identification of human remains by DNA testing methodologies.
Description of human polymorphisms of forensic interest, with special interest in DNA polymorphisms.
Autosomal DNA polymorphisms, mini- and microsatellites (or short tandem repeats, STRs), single nucleotide polymorphisms (SNPs), X and Y chromosome polymorphisms, mtDNA polymorphisms, and any other type of DNA variation with potential forensic applications.
Non-human DNA polymorphisms for crime scene investigation.
Population genetics of human polymorphisms of forensic interest.
Population data, especially from DNA polymorphisms of interest for the solution of forensic problems.
DNA typing methodologies and strategies.
Biostatistical methods in forensic genetics.
Evaluation of DNA evidence in forensic problems (such as paternity or immigration cases, criminal casework, identification), classical and new statistical approaches.
Standards in forensic genetics.
Recommendations of regulatory bodies concerning methods, markers, interpretation or strategies or proposals for procedural or technical standards.
Quality control.
Quality control and quality assurance strategies, proficiency testing for DNA typing methodologies.
Criminal DNA databases.
Technical, legal and statistical issues.
General ethical and legal issues related to forensic genetics.