Targeted multiplexed next generation RNA sequencing assay for tissue source determination of forensic samples

Abstract

The recovery of a DNA profile from the perpetrator or victim in criminal investigations can provide valuable ‘source level’ information for investigators. However, a DNA profile does not reveal the circumstances by which biological material was transferred. Some contextual ‘activity level’ information can be obtained by a determination of the tissue or fluid source of origin of the biological material as it indicates some behavioral activity on behalf of the individual that resulted in its transfer from the body. Here, we seek to improve upon established molecular based methods for body fluid identification (m/miRNA profiling) by developing a targeted multiplexed next generation sequencing assay. The multiplexed biomarker panel includes several highly specific gene targets for each body fluid or tissue with the necessary specificity to definitevely identify all forensically relevant biological fluids and tissues (blood, semen, saliva, vaginal secretions, menstrual blood and skin). We demonstrate the sensitivity of the assay and the ability to identify body fluids in single source and with a limited number of admixed stains. The NGS-RNA body fluid identification panel not only provides ‘activity level’ information for criminal investigations, but will serve as the basis for a future assay which will also permit an association of a DNA profile to a specific body fluid or tissue through the analysis of coding region SNPs within each individual mRNA target. Thus use of this RNA-NGS system may provide forensic scientists with the routine capability of providing some activity level context to a specific individual’s DNA profile.