Verification and evaluation of a miRNA panel for body fluid identification using DNA extracts

Molecular-based approaches for biological source identification are of great interest in the forensic community because of a lack of sensitivity and specificity in current methods. MicroRNAs (miRNAs) have been the subject of many body fluid identification studies due to their robust nature and tissue specificity; however, analysis requires a separate RNA extraction, requiring an additional step in the forensic analysis workflow. The purpose of this project is to build on previous work in our laboratory, wherein we identified a panel of 9 miRNAs that can identify blood, semen, menstrual secretions, vaginal secretions, feces, urine. and saliva. As part of two previously NIJ-funded projects, we conducted a pilot study in which we showed that miRNAs are consistently detectable using several DNA extraction methods commonly utilized in the field for forensic casework. We reported that the miRNA panel for forensic body fluid identification was evaluated using DNA extracts of semen, saliva, blood and menstrual secretions, and was largely concordant with results from samples deriving from RNA extracts. For the proposed project, we will evaluate a larger sample set of DNA extracts and validate the full body fluid panel that was previously validated for RNA extracts using RT-qPCR analysis of DNA extracts. We will perform DNA extractions and query a population sample set of 50 individuals for consistency in differential expression of the mi RNA panel, and therefore body fluid identification. The body fluid samples will vary in age, ethnicity, and gender (where appropriate) of donor. Additionally, 200 mock evidence samples extracted with commonly used DNA extraction methods will be subjected to environmental and chemical insults, and over time in the same individuals to verify our previous findings in DNA extracts. Once validated through this project, a simple RT-qPCR assay can be used with a small portion of the DNA extract to identify the body fluid(s) present in the evidence, with no additional sample use or personnel time in producing a separate RNA extract. Expected technology transition products for this technology will include 1-2 published manuscripts and two to four conference presentations. in addition to the interim and final reports. Note: This project contains a research and/or development component, as defined in applicable law, and complies with Part 200 Uniform Requirements - 2 CFR 200.210(a)(14). CA/NCF