With rapidly improving chemistries and decreasing cost, massively parallel sequencing has incredible potential for forensic investigations. Sequencing forensic short tandem repeats (STRs) can overcome some of the limitations of genotyping by capillary electrophoresis and provides increased statistical significance with backward compatibility to size-based methodologies. The information provided by massively parallel sequencing can be invaluable for the deconvolution and analysis of complex DNA mixtures often obtained from forensic evidence, including items handled by multiple contributors and samples taken from rape cases involving multiple suspects. Additionally, this methodology allows for the analysis of large panels of other forensically relevant DNA markers, such as single nucleotide polymorphisms (SNPs) for identity, phenotype prediction, and ancestry determination. These markers can be used to provide investigative leads and due to their small size, can be exploited for use on highly degraded samples, such as aged evidence or skeletal remains.
Bode Cellmark Forensics, Inc. in collaboration with NexGen Forensic Sciences, LLC tested forensically relevant samples, such as handled documents, gun grips, tool handles, blood, semen, and saliva utilizing several commercially available next-generation forensic STR amplification kits, all specifically designed for use on the Illumina® MiSeq® platform, and a novel Graphical User Interface (GUI) software solution, NexGenID. Under this effort, a method of computational analysis to isolate and identify the unique sequence strings within raw data sets was developed and tested for resolving power. These techniques allowed for the analysis and data interpretation of massively parallel sequencing data of forensic loci of interest, from which the deconvolution of complex DNA mixtures, containing 3 or more donors, was accomplished through the exploitation of SNPs within and immediately flanking STR loci. The advantages of the developed approach lies in its applicability to low copy template samples, relatively rapid turnaround time, and the ability to separate and identify low-level minor contributors to a sample based on the massively parallel nature of the platform. Additionally, the sequence variants analyzed provided an advantage over traditional capillary electrophoresis technologies by adding statistical power to match probabilities, forensic likelihood ratios, and paternity indices. In a panel of 92 individuals comprised of Caucasians, African Americans, Han Chinese, and Mexican Americans, D21S11 demonstrated a match probability of 1 in 23. By analyzing the SNP variants within and surrounding this locus, the match probability increased almost three-fold to 1 in 60. The results of this effort demonstrate the feasibility of complex mixture deconvolution of forensically-relevant STR loci from next-generation sequencing.
Analysis for metagenomics, antimicrobial resistance, and forensics , Gene editing, synthetic genomics, forensics, and biosurveillance
