A comparison of stochastic variation in mixed and unmixed casework and synthetic samples Jo-Anne Bright a, *, Kurt McManus a , SallyAnn Harbison a , Peter Gill b,c , John Buckleton a a ESR, Private Bag 92021, Auckland, New Zealand b Institute of Forensic Medicine, Oslo University, Norway c Centre for Forensic Science, University of Strathclyde, Glasgow, UK 1. Introduction In forensic DNA analysis certain parameters are estimated from empirical data and used in casework interpretation [1–3]. In mixture analysis the variability in the ratio of two peaks of a heterozygote, termed heterozygote balance, Hb, and the variabil- ity in mixture proportion estimate between loci, symbolised D, are often used. There has been some justifiable criticism that the data behind an assessment of this variability is developed from samples, such as mixtures, constructed from pristine DNA [4]. The criticism is that this practice does not adequately mimic casework conditions. Although not explicitly stated in a critique that we can locate, criticism of this type could be extended to include the observation that the heterozygote balance variation is often generated from single source data and then applied to mixtures. 2. Method A previous publication presented a study of the variability in heterozygote balance values for single source crime samples examined at ESR, the provider of forensic services in New Zealand [5]. The same study also presented data on the variability across loci for 36 mixed DNA profiles. These profiles were made from pristine DNA amplified with the Identifiler TM multiplex kit from Applied Biosystems. Casework mixture data was sought to compare with these studies. Casework data suffers from the drawback that the true donors of DNA to the mixture are unknown. In those circumstances of interest for this study there may be ambiguity in the genotype that may be reliably inferred from the electropherogram. An assump- tion that the DNA of the victim is present in a mixture is seldom contentious when the sample source is an intimate sample from that individual. However, the assumption that the DNA of the accused is present is, correctly, strictly avoided by forensic scientists. There are, therefore, considerable barriers, real and psychological to the use of casework data. However, for the purposes of this study such an inference was unavoidable. Casefiles were examined and data collated from electropher- ograms suggesting two person mixtures where the circumstances allowed a reasonable inference about the contributors. The DNA profiles were from a mixture of body fluids extracted using DNA IQ TM (Promega Corporation) method for saliva, bloodstains and trace samples and the Differex TM /DNA IQ TM method for semen/ epithelial mixed stains. All samples were quantified using Quantifiler TM (Applied Biosystems) and 1.5 ng of DNA was targeted for Identifiler TM amplification on a 9700 thermal cycler (Applied Biosystems) with a silver block. Amplified DNA was analysed using a 3130xl capillary electrophoresis instrument and DNA profile data was analysed using GeneMapper TM ID software (Applied Biosystems). Hence the datasets for this paper and the comparison data from pristine DNA mixtures were developed on the same platform. Forensic Science International: Genetics 6 (2012) 180–184 A R T I C L E I N F O Article history: Received 4 June 2010 Received in revised form 29 March 2011 Accepted 1 April 2011 Keywords: Mixed DNA profile Heterozygote balance Mixture proportion A B S T R A C T Understanding the behaviour of mixed DNA profiles is of paramount importance in forensic DNA analysis. Key parameters are those of heterozygote balance and mixture proportion and its variability. These parameters have been previously explored as a function of the average peak height of the active alleles in single source and mixed samples derived from pristine DNA. Here we report a comparison of this data with data obtained from casework samples. This allows an assessment of the difference in the distribution of heterozygote balance between mixed and single source stains and between casework mixtures and synthetic mixtures constructed from pristine DNA. ß 2011 Elsevier Ireland Ltd. All rights reserved. * Corresponding author. E-mail address: jo.bright@esr.cri.nz (J.-A. Bright). Contents lists available at ScienceDirect Forensic Science International: Genetics jou r nal h o mep ag e: w ww .elsevier .co m /loc ate/fs ig 1872-4973/$ – see front matter ß 2011 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.fsigen.2011.04.010