Biological traces found at crime scenes can be analyzed to genetically identify the donor(s) but also to determine the body fluid composition of the stain. The latter can be accomplished with high specificity by mRNA profiling. In some mixed body fluid samples, it might be of probative value to directly associate each body fluid with each of the DNA donors. Associating a DNA profile to the contributing body fluid is not often possible, with the exception of simple binary mixtures that contain male and female specific body fluids. However genomic information is transferred from DNA to mRNA, and RNA-cSNPs (coding region SNPs) should be identifiable by MPS methods. Previously, we have performed proof-of-concept studies using a prototype Illumina MiSeq MPS cSNP assay to demonstrate the usefulness of this approach. Here, we have continued our work and introduce an MPS assay on the Ion S5 system comprising a set of 21 cSNPs in body fluid specific mRNA transcripts (7 blood (3 genes); 8 semen (4 genes); 6 saliva (4 genes)). Combined with optimized primer sets for body fluid identification, the assay can identify all forensically relevant body fluids and skin as well as differentiating blood, semen and saliva transcripts from different individuals.
The assay has been evaluated with numerous donors of each body fluid to evaluate the specificity and the discriminatory power of the cSNPs. The findings are promising as we were able to associate donors with body fluids in mixtures of body fluids. However, more cSNPs are needed to improve the discriminatory power, particularly for vaginal secretions and menstrual blood transcripts. Assigning body fluids to DNA donors is a realistic possibility with the continued development of MPS cSNP assays.