How it got there: associating individual DNA profiles with specific body fluids in mixtures using targeted digital gene expression and RNA-SNP identification

As submitted by the proposer: The recovery of a DNA profile from biological evidence does not reveal the circumstances by which biological material was transferred. A determination of the tissue or body fluid origin of this material would provide some indication of the activity preceding the deposition of the DNA profile. Traditional biochemical methods do not exist for the identification of all forensically relevant biological fluids and tissues. Currently mRNA profiling is the only available fully validated molecular based method for comprehensive body fluid identification. Even with this approach, it is not possible to associate the component DNA profiles with specific body fluids, a requirement in order to obtain objective 'activity level' information in criminal investigations. Therefore, this project seeks to improve upon mRNA profiling-based body fluid identification by the development of a targeted multiplexed next generation RNA-sequencing system capable of providing a definitive link between a DNA profile and a particular fluid or tissue type, a novel piece of probative information that cannot be obtained using any other body fluid identification method (e.g. CE or qPCR analysis, epigenetics, proteomics). We will design a highly multiplexed 78-marker panel that will include the analysis of numerous gene targets for each fluid or tissue. The primer sets will be designed so that, in addition to digital gene expression data, genotype data from coding region SNPs (or RNA-SNPs) within each mRNA target will be obtained simultaneously. The inclusion of -70 polymorphic RNA-SNPs should result in a high probability of discrimination between two random individuals, thus establishing a direct and definitive link between a DNA profile and a particular fluid or tissue in admixed samples. After initial validation of the developed assay, it will be transferred to the Arizona Department of Public Safety (AZDPS) crime laboratory who will perform a full casework evaluation and validation, including blind testing, to further test the accuracy and reliability of the assay. The project team (UCF, AZDPS and lllumina Corporation) represents an ideal opportunity to support the forensic community through the collaborative effort of the forensic research, casework and industry sectors. We envision the possibility that the results of this study could form the basis of a commercial product that would facilitate the transfer of the body fluid identification methods to casework laboratories. The results of this study will be presented to the forensic community through comprehensive technical reports, peer-reviewed articles and presentations at domestic and international scientific conferences. ca/ncf