FaSTR™ DNA makes analysis of profiles generated from the new PowerPlex® 35GY 8-dye kit on the Spectrum CE instrument faster and easier through a variety of innovative features including:

Customizable rules

• FaSTR™ DNA provides fully customizable rules for handling of common analysis artefacts reducing user interaction and analysis time required.
• Settings can be configured for rules to handle off-ladder peaks, split or quenched peaks, N or shoulder peaks, pull-up, cross-channel artefacts (artefacts that occur in the same location across multiple dye channels), and peaks with poor morphology.

Automatic detection of known artefacts

Known artefacts are published either by the kit manufacturer and/or documented by a laboratory.

• The ability to automatically filter or flag artefacts in fixed or relative positions as well as artefacts occurring in the same location across all dye channels has also been incorporated in the FaSTR™ DNA software.
• Keyboard shortcut keys for quick application of most commonly used interactions are available and a PDF of these can be printed from within the software.

Customizable stutter filtering

• FaSTR™ DNA detects an unrestricted number of stutter types in single- or composite stutter positions using a range of stutter model options to determine the expected stutter ratio.
• Levels for automatic stutter filtering can be customized by locus or by allele, including use of stutter regression and exceptions files which align with the files used in STRmix™ to model stutter [1, 2]. Adjustments to stutter filter (and flag) levels can be automated for where composite stutters are involved. This negates the need for manual calculations and review.

Reviewing peaks and samples

• A simple color-coded system of flagging samples requiring user review has been implemented within FaSTR™ DNA.
• Peaks that require analyst review have highlighted peak labels to catch the analyst’s attention.
• A peaks table is provided for all peaks detected within a sample and the rules applied to each peak are detailed in the table. Within the table, a separate tab shows just the peaks that need analyst attention.
• Peaks that have been edited manually by the analyst are recorded in a separate list of changes.
• Removed peaks remain selectable and can be reviewed with all peak details retained.
• All peak data, regardless of peak status, can be exported.

Number of contributors (NoC) estimation

• Automated estimation of NoC using a decision tree [3] or maximum number of alleles calculation is available within the software.
• Analysts can also assign their own estimate of NoC.
• The NoC assigned within FaSTR™ DNA is used when integrating with STRmix™ to seamlessly transition samples from analysis to interpretation.

Project Review

• FaSTR™ DNA includes a project review module allowing projects of corresponding samples analyzed independently to be directly compared with the differences highlighted in the analysis graph.
• The project reviewer can resolve any differences between two analysts or between one analysis and the raw analyzed project and create a final project ready for export.
• The project review module can also be used as a training tool as it allows analysts to compare their own analysis decisions against an exemplar analysis of the same sample set (or the raw analyzed project). Where differences are observed, the trainee can clearly articulate the reasoning behind their conclusions if needed, thus strengthening their decision-making skills and their ability to clearly articulate and defend those decisions.

Comparison module

• FaSTR™ DNA includes a comparison module that allows sample-to-sample comparison within a project, as well as comparison of samples within the project to a saved database. This can be used to investigate sample-to-sample correspondences, as well as correspondence to, for example, a staff elimination database.

Additional rules

• FaSTR™ DNA can automatically check for control sample and quality marker concordance and alert the user if any issues are detected.
• From FaSTR™ DNA 1.3, visible indication of pull-up can be displayed on the electropherogram (EPG).

The latest release, FaSTR™ DNA 1.3, provides the ability to automatically assign NoC separately for the autosomal as well as the Y-STR loci contained with the PowerPlex® 35GY System. Y-NoC estimation is based around a series of customizable rules and optional sample warnings allowing users to automatically estimate Y-NoC in a manner aligned with laboratory specific manual Y-NoC processes.

FaSTR™ DNA 1.3 also allows users to add notes directly to epgs on screen and optionally include these in epg reports. Samples analyzed as part of a large project can be pulled out into their own analysis project providing an improved workflow for case management.

The core algorithms within FaSTR™ DNA are adapted from the process described by Goor et al. [4], and the developmental validation of FaSTR™ DNA has been published [5].

The use of FaSTR™ DNA for analysis of Promega PowerPlex® 35GY 8-dye data on the Spectrum CE System was validated and implemented for NHSPFL following the FBI Quality Assurance Standards (QAS). This included the development of a set of laboratory specific stutter models and corresponding stutter filters for use with both FaSTR™ DNA and STRmix™; custom methods for use for both casework and single source profiles; and validation of a NoC decision tree for autosomal NoC estimation. The performance metrics of autosomal NoC estimation using a decision tree available in FaSTR™ DNA were assessed using a set of 98 laboratory designed mixed DNA profiles.

The results of this performance check are summarized in Table 1 below.

Instances of over- or under-estimation of NoC by the decision tree were investigated further. Mis-classifications were due to the presence of elevated stutter, drop-in, peak height imbalance, and the use of sub-AT data by the analyst when manually determining NoC. All outcomes and reasoning were intuitive given the observed profiles.

The New Hampshire State Police Forensic Laboratory (NHSPFL) is the first laboratory in the world to implement FaSTR™ DNA and STRmix™ probabilistic genotyping software into operational casework using the Promega PowerPlex® 35GY 8-dye kit on the Spectrum CE System. This was an example of a successful collaboration between the STRmix team, Promega and NHSPFL. NHSPFL reports significant efficiency gains to their workflow with reduced turnaround time for profile analysis with the numerous FaSTR™ DNA tools and features as well as lower fatigue from the customizable ergonomic visual design of the software. Further, the integration of the FaSTR™ DNA and STRmix™ software has lessened the logistic complexity of interpretation set up allowing for a streamlined approach from analysis to interpretation.