Precision-ID

The Applied Biosystems  Precision ID NGS System for human identification can help you solve tough cases by getting more information from your challenging samples. Now you have help moving those unknown samples from storage to investigative leads. Adopting next-generation sequencing (NGS) for forensic DNA analysis in your laboratory is simpler than ever when you combine the Ion Chef System and Ion GeneStudio S5 Systems with optimized Precision ID library preparation, template preparation and sequencing kits, and forensically relevant panels.

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Let the evidence speak for itself. You can choose from a suite of targeted Precision ID panels for the NGS solution for your cases. Mitochondrial DNA (mtDNA) from samples recovered at mass disasters and from other unidentified remains can be routinely analyzed to assist in human identification, and help establish links to related family members. Short tandem repeat (STR) panels provide high-resolution genotyping of STRs for analysis of complex mixtures. Ancestry-informative or phenotypic single nucleotide polymorphism (SNP) analysis may help generate investigative leads when suspects are unknown. Identity-testing SNPs can associate a degraded crime scene sample to a known reference when partial results are obtained with autosomal STR analysis.

With as little as 125 pg of DNA input, you can process unsolved and missing persons cases with NGS in your lab today, with as few as 5 pipetting steps and 45 minutes of hands-on time.

Solutions for forensic DNA analysis
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Applied Biosystems Precision ID panels enable analysis of up to hundreds of forensically targeted markers such as the mtDNA genome, STRs, or SNPs.

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The Ion Chef System simplifies the workflow for targeted sequencing, providing a walk-away solution for automated library preparation, template preparation, and chip loading.

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The Ion GeneStudio S5 Systems provide the simplest DNA-to-data workflow for targeted sequencing—so you can spend less time doing repetitive lab work.

Simplest targeted NGS workflow

Construct library
Hands-on time: 15 minutes
Total time: 7 hours

Prepare template
Hands-on time: 15 minutes
Total time: 11 hours

Run sequence
Hands-on time: <15 minutes
Total time: as little as 2 hours

Analyze data
Total time: as little
as 5 hours

 

Precision ID panels

 

Ion Chef System

 

 

Ion GeneStudio S5 System

 

Converge Software
for mtDNA, STR
and SNP analysis

Precision ID panels for human identification

Number of samples per run

Category

Panel
Ion S5 Chips
    Ion 510 Chip
Ion 520 Chip
Ion 530 Chip
mtDNA
Precision ID mtDNA Whole Genome Panel
 
25
32
Precision ID mtDNA Control Region Panel
37
56
 
STR
Precision ID GlobalFiler NGS STR Panels
 
16
32
SNP
Precision ID Ancestry Panel
48
72
362
Precision ID Identity Panel
54
81
384

Recommendations are based on in-house determination of the number of samples that can be multiplexed while still achieving a minimum coverage of 100x for 97% of SNP amplicons for SNP panels and all mtDNA amplicons.  Individual lab results may vary depending on workflow used and customer requirements.

Identify missing persons or analyze remains
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In forensic casework, mitochondrial DNA (mtDNA) is useful in the context of challenging samples that fail to produce an autosomal STR profile due to their high copy number per cell. When samples such as teeth, bones, and hairs without a follicle (root) are collected from crime scenes, missing-person remains, or disaster areas, even amplicon coverage and minimal strand bias are key factors that contribute to high-quality genetic results for successful identification of human remains.

The Applied Biosystems Precision ID mtDNA Whole Genome Panel is an innovative approach to mitochondrial DNA sequencing specifically developed for forensic applications. This mtDNA tiling approach, using amplicons that are only 163 bp average length, assists with obtaining optimal mitochondrial genome (mtGenome) coverage from highly compromised, degraded samples such as hair shafts, teeth, and bones.

The Applied Biosystems Precision ID mtDNA Control Region Panel is based on the same tiling approach used in the Precision ID mtDNA Whole Genome Panel. This targeted control region panel spans the entire 1.2-kb control region, which encompasses HV-I, HV-II, and HV-III, with the same optimal small amplicon design ideal for performance with degraded forensic samples. In addition, the panel design leverages primer degeneracy in known variable regions to help ensure robust performance across diverse population samples.

Analysis of the mtGenome can be challenging due to complex alignments, the presence mtDNA heteroplasmy, and insertions and deletions present throughout the genome that may impact the accuracy of variant calling. The Applied Biosystems Converge Software NGS Data Analysis module automates mitochondrial DNA analysis leveraging optimized base calling, alignment and quality filtering algorithms specific for the Precision ID mt DNA panels. These solutions gives forensic DNA laboratories the flexibility to detect variation within noncoding control region sequences or take full advantage of the genetic diversity with analysis of full mtGenome sequence data using either the Precision ID mtDNA Control Region Panel or Precision ID mtDNA Whole Genome Panel using as little as 2 pg of DNA.

Analyze DNA mixtures more efficiently

The Applied Biosystems Precision ID GlobalFiler NGS STR Panel v2 includes 36 markers, including the same 21 autosomal STRs along with Y markers and amelogenin sex markers found in the GlobalFiler PCR Amplification Kit, as well as 14 additional informative markers for forensic analysis. When analyzed with NGS, these markers reveal more alleles (74%) than identified with traditional CE analysis due to sequence diversity within CE alleles of the same size. These isometric alleles can be used to help resolve complex profiles and mixtures.

With as little as 125 pg of DNA input, you can quickly and easily analyze these commonly used forensic markers in less than 2 days.

The Precision ID GlobalFiler NGS STR Panel provides both allele number and base sequence for each repeat structure, offering enhanced mixture resolution and increased Probability of Identity (PI) estimates.

Generate more investigative leads
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Precision ID Ancestry Panel
The Applied Biosystems  Precision ID Ancestry Panel can provide you with biogeographic ancestry information and guide your investigation process. Utilizing Ion AmpliSeq technology, the Precision ID Ancestry Panel helps to enable simple and fast target selection of hundreds of single nucleotide polymorphisms (SNPs) using multiplex PCR. This ready-to-use panel consists of 165 autosomal markers. A total of 55 of these markers were selected based on a poster presented by Dr. Kenneth Kidd, and 123 markers were selected based on a publication by Dr. Michael Seldin. This sensitive panel, with average amplicon sizes of <130 bases, is optimized for degraded DNA samples and uses as little as 125 pg of input DNA.

The Converge Software NGS Data Analysis module contains parameters for analyzing the Precision ID Ancestry Panel, generating admixture predictions as well as the population likelihood from 151 ancestry-informative markers that cover seven continental populations.


Analyze highly degraded or trace DNA

Precision ID Identity Panel

Single-nucleotide polymorphisms (SNPs) provide a variety of new, important information for human identification purposes, in addition to short tandem repeats (STRs). Due to the shorter amplicons required for sequencing, SNPs enable high recovery of information from degraded samples—such as from mass disaster victims.

The   Applied Biosystems Precision ID Identity Panel includes 124 autosomal markers, and provides discrimination of individuals similar to STR genotype match probabilities used by forensic analysts (between 1 x 10-31 and 6 x 10-35). The high discrimination power is achieved by using 34 upper Y-clade SNPs and 90 autosomal SNPs that have high heterozygosity and low Fixation Index (Fst), described in publications by Kenneth Kidd of Yale University and the SNP for ID Consortium.

The Converge Software NGS Data Analysis module contains parameters for analyzing the Precision ID Identity Panel, generating random match probabilities (RMPs) from the 1000 Genomes Project dataset using 85 unlinked identity SNPs, as well as Y haplogroup determination from the 34 upper Y-clade SNPs.

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The Applied Biosystems Precision ID Library Kits are specifically designed for rapid generation of targeted sequencing libraries from Precision ID panels. The kits are built on Ion AmpliSeq chemistry, which enables scalable multiplex PCR reactions from tens to thousands of amplicons in a single well using as little as 125 pg of starting DNA. These kits also enable the preparation of barcoded libraries using Ion Xpress Barcode Adapters 1-96 Kits or the IonCode Barcode Adapters 1-384 Kit. The barcoded libraries can be combined and loaded onto a single Ion chip to minimize the sequencing run time and costs, and allow for accurate sample-to-sample comparison. Precision ID Library Kits are for use with manual procedures or with high-throughput robotics platforms.

The Precision ID DL8 Kit is designed for use with the Ion Chef System for automation of Ion AmpliSeq library construction. Leveraging the automation capabilities of the Ion Chef System, this kit enables the reproducible preparation of Precision ID amplicon libraries, with less than 15 minutes of hands-on time and only 3 liquid transfer steps. Compatible with 1- and 2-pool panel designs, the kit automates the preparation of libraries from 8 samples in a single run, delivering a single tube of pooled libraries ready for downstream templating and sequencing.

The Ion S5 Precision ID Chef & Sequencing Kit is fully optimized for templating and sequencing of all available HID NGS panels (STR, SNPs, and mitchondrial DNA) on the Ion GeneStudio S5 Systems. The kit is available in two formats: two runs per initialization for higher-throughput labs and one run per initialization for less frequent analyses.

Automate library and template preparation with the Ion Chef System

Save time at the bench, help improve your laboratory’s productivity, and standardize your results. The Ion Chef System for human identification performs the repetitive lab work without the need for complex robotics and confusing scripts. Paired with Ion Torrent next-generation sequencing platforms, the Ion Chef System automates Precision ID library preparation, template generation, and chip loading with push-button simplicity—enabling DNA to data in as little as 45 minutes of hands-on time.

Ion Chef System templating and chip loading workflow
1 Library preparation
 
 
  • Load reagents & consumables
  • Load library samples
  • Set run parameters
 
2 Automated template preparation and chip loading
  • Library amplification
  • ISP recovery & enrichment
  • Chip loading
 
3 Sequencing
 
 
Ion PGM, Ion S5

Run sequence on:

  • Ion S5 GeneStudio System

 

Suitable for scientists and researchers in forensic human identification, the Ion Chef System offers automated workflows that help enable gains in laboratory efficiency. Unleash the power of the Precision ID NGS System with Ion Chef System automation, paired with Ion AmpliSeq technology which is used in the Precision ID Panels for human identification.

  • Automates library generation, equalization, and pooling for 1- and 2-pool designs for up to 8 samples in a single instrument run
  • Operates with a very low inter-run cross-contamination rate of <0.01%
  • Helps reduce sources of variability for forensic scientists of any experience level
  • Supports all Ion Torrent semiconductor chips and sequencing chemistries

Comparison of reproducibility across several next-generation sequencing metrics (number of reads, base coverage and loading efficiency) for Ion libraries processed (templating and chip loading) using the Ion Chef and Ion OneTouch 2 Systems. While both systems exhibit excellent reproducibility, the Ion Chef System demonstrates lower coefficient of variation (CV) % values across all metrics.

Fast, flexible, and simple next-generation sequencing for forensics with the Ion GeneStudio S5 Systems
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Want to sequence SNP panels on Monday, mtDNA panels on Wednesday, and STR panels on Thursday? The Ion GeneStudio S5 Systems, an integral part of the Precision ID NGS System, let you leverage a single benchtop instrument that scales to your application and throughput needs. The Ion GeneStudio S5 Systems provide the simplest DNA-to-data workflow for targeted sequencing with industry-leading speed and affordability, and the flexibility to multiplex and optimize the number of samples and panels on a single chip.

Simplicity: ready, set, sequence

  • Less than 15 minutes of sequencer hands-on time
  • Less than 45 minutes of hands-on time for a DNA-to-data targeted sequencing workflow

Speed: because every hour counts

  • <2.5 hours for a sequencing run
  • From DNA to data in less than 2 days with an overnight Ion Chef templating run
  Ion GeneStudio S5 System Ion GeneStudio S5 Plus System Ion GeneStudio S5 Prime System
    Ion 510 Chip Ion 520 Chip Ion 530 Chip Ion 510 Chip Ion 520 Chip Ion 530 Chip Ion 510 Chip Ion 520 Chip Ion 530 Chip
Reads   2-3 million 4-6 million 15-20 million 2-3 million 4-6 million 15-20 million 2-3 million 4-6 million 15-20 million
Turnaround time (sequencing + analysis) mtDNA whole genome panel - 7.5 hr 10.5 hr - 3.5 hr 5 hr - 3 hr 4 hr
mtDNA control regions panel 4.5 hr 7.5 hr - 3 hr 3.5 hr - 3 hr 3 hr -
Ancestry panel 4.5 hr 7.5 hr 10.5 hr 3 hr 3.5 hr 5 hr 3 hr 3 hr 4 hr
Identity panel 4.5 hr 7.5 hr 10.5 hr 3 hr 3.5 hr 5 hr 3 hr 3 hr 4 hr
STR panel - 12.5 hr 22.5 hr - 4.5 hr 5.5 hr - 4 hr 5 hr
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Small sample input: because every sample matters

  • As little as 125 pg DNA needed to generate human identification profiles

Scalability: single sequencer, multiple applications

  • Analyze SNPs, mtDNA, STRs and mRNA targets
  • Multiple chip formats and read lengths to match your throughput needs 
  • Flexible multiplexing of samples and panels on a single chip
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Simple data analysis and storage

  • Plan, monitor, track, and analyze your runs in Torrent Suite Software
  • Integrate, annotate, and interpret variants

Service and support

  • More than 3,700 global sales, service, and technical support specialists are available to assist you in person, by phone, or online—including our Human Identification Professional Services (HPS) team that can help you successfully navigate the validation process required to bring new technologies into your lab.

 

New to next-generation sequencing? Watch the demos for each step of the Ion Torrent next-generation sequencing workflow.

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Converge Software

An all-in-one modular enterprise platform from Thermo Fisher Scientific, integrates forensic DNA data management and analysis into a single software package designed to increase the efficiency of forensic DNA laboratories. With the Converge NGS analysis module, laboratories are now able to examine the mitochondrial genome to identify remains when there is poor quality or no autosomal DNA available for analysis, STR markers to help determine the number of contributors in a mixture analysis, and targeted and forensically relevant single nucleotide polymorphism (SNP) markers to help generate investigative leads. Additionally, full auditing functionality is included for chain-of-custody requirements.

mtDNA analysis

Analysis of the mtGenome can be challenging due to complex alignments, the presence mtDNA heteroplasmy, and insertions and deletions present throughout the genome that may impact the accuracy of variant calling. NGS reads from the BAM files are first mapped to nodes in PhyloTree and then realigned using a custom Smith–Waterman alignment algorithm that integrates PhyloTree and EMPOP information into the scoring function. Variants are called with reference to the rCRS. Additionally, the closest haplogroup is calculated, and variants are evaluated based on their occurrence in the haplogroup as well as other general metrics including frequency, strand bias, and coverage. Variants can be viewed in a variety of formats, including a circular plot, linear view or grid.

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STR analysis

With an interface similar to that of Applied Biosystems GeneMapper ID-X Software, you will be able to quickly evaluate sequencing data using familiar process quality values (PQV) and flags such as allele number (AN), off-ladder allele (OL), peak height ratio (PHR), below stochastic threshold (BST), and control concordance (CC) (use figure). Preconfigured analysis settings are provided within the NGS module and may be modified by the laboratory as needed.

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SNP analysis

Converge Software SNP analysis provides a variety of metrics to monitor sequencing quality, including coverage of aligned reads to a hotspot, strand bias, number of reads containing each base at the hotspot, genotype call and quality, and major allele frequency. Tertiary ancestry analysis consists of generating an estimation of admixture prediction and population likelihoods (based on algorithms from Ken Kidd). Identity analysis consists of calculating Random Match Probability (RMP) based on genotype frequencies generated from 1000 Genomes data as well as a Y haplogroup prediction.

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Adapting next-generation sequencing (NGS) for forensics is not a paradigm shift —Dr. Bruce Budowle, University of North Texas Health Science Center
 Watch now ›

Introducing mitochondrial DNA sequencing by NGS technology —Dr. Walther Parson, Institute of Legal Medicine at Innsbruck Medical University
 Watch now ›

Accelerate your forensic DNA analyses with Ion Torrent NGS automation —Joseph Chang, Thermo Fisher Scientific
 Watch now ›

Mitochondrial DNA analysis

STR analysis

Ancestry SNP analysis

Identity SNP analysis

Customer videos

Massively Parallel Sequencing of mtDNA in a Casework Laboratory

How does mitochondrial DNA (mtDNA) analysis play a role in making identifications in missing persons cases? What are the similarities and differences between MPS and Sanger sequencing workflows for mtDNA analysis? Jennifer Churchill, PhD, Research Assistant Professor at the University of North Texas, provides guidance for bringing mtDNA analysis into casework operations.

Mitochondrial DNA analysis in human identification

Imagine that you have a small, aged bone fragment or charred remains after a fire, a tooth that has been buried and exposed to the environment for years to decades, or possibly a single hair shaft. How would you even begin to make an identification to reconnect these remains with family members? Walther Parson, PhD from the Institute of Legal Medicine, Innsbruck Medical University discusses a number of cases his lab has worked on, using mtDNA analysis to make critical identifications.

MPS of STRs in forensic DNA analysis

Forensic scientists need to differentiate genetic profiles from complex forensic DNA mixtures, such as those commonly associated with crime scenes. But complicating factors, such as DNA degradation, mixtures and/or insufficient starting material, may prevent an estimated 30–40% of samples from generating conclusive results when using traditional capillary electrophoresis–based approaches. Now, by using the Applied Biosystems Precision ID GlobalFiler NGS STR Panel v2 and Converge Software 2.0, labs can effectively retrieve more information from these mixed, degraded or limited DNA samples.

Next-generation sequencing for forensic DNA analysis

Next-generation sequencing and the ability to multiplex many markers for a single sample has gained acceptance within the forensic community. There are a number of SNP classifications that are currently being investigated, including identity SNPs, phenotypic SNPs, and biogeographic ancestry SNPs. Executive Director Soizic Le-Gunier discusses how Institute Genetics Nantes Atlantique (IGNA) uses the various technology options, and how SNP genotyping will have an impact in the future.

References