Forensic DNA Analysis of Challenging Samples

The Applied Biosystems AutoMate Express Forensic DNA Extraction System is an easy-to-use, robust benchtop instrument that offers maximum flexibility and utilizes the Applied Biosystems PrepFiler Express and PrepFiler Express BTA (bone, tooth and adhesive) chemistries packaged in prefilled, foil-sealed cartridges. The ease of instrument setup, minimal handling of sample tubes, and short run time make the AutoMate Express system an ideal instrument for analysts to quickly perform unscheduled extractions for expedited cases. More importantly, the AutoMate Express system offers the greatest range of elution volumes (from 20 µl to 250 µl) and maximizes the ability to obtain interpretable STR results. 

The quantification step of the casework workflow is the key to determining how a sample will be processed downstream. There are many factors that determine the success of the short tandem repeat (STR) profile. Our advanced quantification instrumentation, kits and software enable forensics labs to achieve accurate results in less time for even the most challenging casework samples. The value of DNA quantification includes:

• Improves amplification efficiency
• Results in fewer off-scale, over-amplified, or under-amplified samples
• Gives normalized profiles, resulting in reliable data interpretation
• Designed to reduce the need for downstream reanalysis, which enables long-term cost savings
• Provides key sample information to enable forensic workflow decisions
• Enables conservation of sample through targeted downstream assay decisions

The Applied Biosystems Quantifiler HP and Quantifiler Trio DNA Quantification Kits enable forensics laboratories to simultaneously obtain a quantitative and qualitative assessment of total human DNA in a single, highly sensitive real-time PCR reaction.

Dr. Sheree Hughes-Stamm , Sam Houston State University, Houston, Texas

Applied Biosystems HID Real-Time PCR Analysis Software v1.3 provides the ability to analyze the data with a virtual standard curve that is user defined. In addition, the software includes the quantification, STR setup features, and STR data quality review functionality that integrate and streamline the quantification and STR analysis steps.

As global forensic DNA databases rapidly expand, so does the need for more discriminating STR multiplexes that can maximize loci overlap. That’s why Applied Biosystems GlobalFiler STR kits incorporate the most commonly used loci—all in a single multiplex reaction. GlobalFiler kits contain all of the STR loci commonly used in major global databases, including all markers recommended for inclusion by the CODIS Core Loci Working Group and those markers commonly used in Europe. The multiplex selection of markers reduces the risk of adventitious matches while enabling more effective cross-border data sharing.

The GlobalFiler kit has been optimized to maximize the quality and quantity of genotyping results, even with the most challenging samples. The robust master mix, 10 mini-STR markers, and expanded DNA input range generate more information from the most difficult samples—those that are highly degraded and/or inhibited.

Dr. Antonio Alonso, National Institute of Toxicology and Forensic Sciences, Madrid Spain

The Applied Biosystems NGM Detect PCR Amplification Kit helps minimize the size of highly discriminating markers, such as SE33. Designed specifically for compatibility with European database requirements, this kit offers superior sensitivity and provides an alternate configuration to the well-established NGM SElect PCR Amplification Kit format, to help maximize information recovery even from degraded casework samples. The NGM Detect kit  helps to enable positioning of markers with the highest representation in databases to be prioritized, to enhance recovery of searchable information from casework samples. An improved master mix formulation coupled with a capacity for higher sample volume in the reaction (15 μL vs. 10 μL DNA) helps to enable high recovery of allelic data.

Y-STR loci can be valuable in forensic investigations because the Y chromosome can be used to trace male lineages due to its unique genetic inheritance from father to sons. Y-STR loci have been serving as a supplementary tool for relationship testing, ancestry identification, familial searching, missing persons and mass disaster victim identification, and especially sexual assault cases to exclude the potential male contributors or further confirm the identity.

Forensic casework, missing persons and mass disaster victim identification in particular pose significant challenges to the analyst in terms of sample quality and quantity. The Applied Biosystems Yfiler Plus kit can get more information with its 27 Y-STR markers, including 7 rapidly mutating Y-STR loci and 11 mini-STR (<220bp), from low quantity or low quality casework samples with its high sensitivity and improved inhibitor resistance. 

Dr. Franz Neuhuber, University of Salzburg, Austria

For analyzing DNA mixtures, the Applied Biosystems Precision ID GlobalFiler NGS STR Panel v2 includes the same 21 autosomal STRs along with Y markers and amelogenin sex markers found in the GlobalFiler PCR Amplification Kit. Instead of using SE-33, this panel includes additional multi-allelic STR markers to aid in mixture resolution for complex casework samples. Using a small sample input of as little as 125 pg of DNA with this targeted forensic marker panel helps to enable you to go from sample extraction to genotyping in less than 2 days. The Precision ID GlobalFiler NGS STR solution provides information about STR allele calls, STR sequence motifs, known SNPs in flanking regions, and isometric heterozygotes (alleles of the same fragment length but containing different sequences).

Dr. Pedro Barrio, National Institute of Toxicology and Forensic Sciences, Madrid Spain

Generate more investigative leads with the Applied Biosystems Precision ID Ancestry Panel, which includes 165 autosomal markers (SNPs) that can provide you with biogeographic ancestry information and guide your investigative process. With small amplicon sizes (<130 bp), this panel is also optimized for degraded DNA samples. Using a small sample input of as little as 1 ng of DNA, you can go from sample extraction to genotyping in a little over a day.

Analyze degraded or trace DNA with the Applied Biosystems Precision ID Identity Panel, comprised of 124 markers and providing discrimination of individuals similar to STR genotype match probabilities used by forensic analysts. This high discrimination power is achieved by using 34 upper Y-clade SNPs and 90 autosomal SNPs. Due to the shorter amplicons required for sequencing, SNPs enable high recovery of information from challenging samples such as samples from mass disasters, low–copy number samples, or low-quality samples.

Dr. Walther Parson, Innsbruck Medical University, Austria

Analyze evidence from missing persons or remains from disaster victims with the Applied Biosystems Precision ID mtDNA Whole Genome Panel, an innovative approach to mtDNA sequencing specifically developed for forensic applications. This mtDNA tiling approach includes two primer pools of 81 small amplicons in each pool, with an average size of 163 bp, to assist with obtaining optimal mitochondrial genome coverage data from highly compromised, degraded samples such as hair shafts, teeth and bones.

Additionally, the Applied Biosystems Precision ID mtDNA Control Region Panel is based on the same mtDNA tiling approach used in the Precision ID mtDNA Whole Genome Panel. This targeted control region panel contains two smaller sets of primer pools with 7 amplicons in each pool that span the 1.2-kb control region, which encompasses hypervariable (HV) regions I, II and III, with the same optimal coverage for degraded forensic samples.