Inferring the Number of Contributors to Complex DNA Mixtures Using Three Methods: Exploring the Limits of Low‐Template DNA Interpretation

Abstract

In forensic DNA casework, the interpretation of an evidentiary profile may be dependent upon the assumption on the number of individuals from whom the evidence arose. Three methods of inferring the number of contributors-NOCIt, maximum likelihood estimator, and maximum allele count, were evaluated using 100 test samples consisting of one to five contributors and 0.5-0.016 ng template DNA amplified with Identifiler Plus and PowerPlex 16 HS. Results indicate that NOCIt was the most accurate method of the three, requiring 0.07 ng template DNA from any one contributor to consistently estimate the true number of contributors. Additionally, NOCIt returned repeatable results for 91% of samples analyzed in quintuplicate, while 50 single-source standards proved sufficient to calibrate the software. The data indicate that computational methods that employ a quantitative, probabilistic approach provide improved accuracy and additional pertinent information such as the uncertainty associated with the inferred number of contributors.

A review of trace “Touch DNA” deposits: Variability factors and an exploration of cellular composition

“…not transferred from elsewhere on the body) nucleated cells may nonetheless be present on people's hands providing touch DNA. STR typing is routinely sensitive enough to detect 0.200 ng or less, (equivalent to approximately 30 nucleated cells [132]) and current methods detect profiles from almost single-cell levels (0.008-0.010 ng) [133,134,135]. Consequently, even a limited population of nucleated cells, possibly originating from eccrine sweat ducts and subsequently brought to the hands' surface, could be a contributing factor in touch deposit DNA.…”

Exploring new short tandem repeat markers for DNA mixture deconvolution

“…Of these mixture samples, 33% of samples had at least two contributors, 9% had at least three contributors and 1% were comprised of four or more contributors ( Figure 7) (Gross, 2008). Since 2008, there are indications that the number of DNA samples presenting as three or more person mixtures has continued to rise, resulting in increasingly complex interpretation of already challenging evidence (Alfonse, Tejada, Swaminathan, Lun, & Grgicak, 2016;Isaacson et al, 2015).…”

Developmental validation of PACE™: Automated artifact identification and contributor estimation for use with GlobalFiler™ and PowerPlex® fusion 6c generated data

“…Samples within this set included 152 different DNA template amounts ranging from 3.0 pg to 5.1 ng and 49 different ratios of contributors. Samples were run on five Applied Biosystems 3500 Genetic Analyzers using 15, 20 and 24 s injections at 1 kV and three Applied Biosystems 3100 series Genetic Analyzers (Thermo Fisher Scientific Inc.) using 3,5,10,11,12,15,20 and 30 s injections times with a 3 kV injection voltage (Table 1).…”

“…Sample 1426, a 2-contributor sample classified as 4+-contributor (Supplementary information, Fig. 1S), was a distinct misclassification; it was amplified using 0.375 ng of template DNA but was subjected to DNase I degradation (24mU enzyme) [3]. This led to low peak heights, with 57.8% of the peaks falling below 100RFU and peak height imbalance that typically indicates greater than two contributors.…”

“…This need helped spur the development of computationally intensive interpretational tools that can probabilistically assess the number of contributors in a DNA mixture. These include NOCit [3], which uses a Monte Carlo sampling algorithm to estimate the posterior probability on n given an electrophoresis profile, and PACE [4], which uses machine learning to estimate the optimal decision boundaries in feature space between the possible values of n, and subsequently constructs a predictive model that estimates n for a given profile.…”