RNA‐based approaches for body fluid identification in forensic science

Abstract In forensic science, the identification of body fluids present in a stain can aid in the reconstruction of crime scenes and support activity‐level information in some cases. RNA‐based methods for body fluid identification address some limitations associated with conventional testing, including a lack of specificity and sample destruction. mRNA profiling by endpoint reverse transcription polymerase chain reaction (RT‐PCR) is a confirmatory RNA method currently used by some forensic laboratories in casework due to the availability of capillary electrophoresis instruments. This method is highly specific and reliable when the RNA input is within the optimal range. When RNA input is below the limit of detection or too high, then false negatives and false positives/cross reactivity can occur, respectively. Real‐time quantitative PCR (qPCR) is quantitative, more sensitive, and efficient than endpoint PCR, and does not require post‐PCR processing. A reverse transcription (RT) step may also be included in qPCR (RT‐qPCR), enabling mRNA quantification. However, there are concerns regarding the lack of consistency and standardization of RT‐qPCR methodology and reporting of results. Additionally, the number of currently available fluorophores somewhat reduces multiplexing ability. More recently, approaches involving Piwi‐interacting biomarkers, NanoString barcodes, and RNA sequencing have been developed. This review will cover the current and developing RNA‐based techniques for forensic body fluid identification, focusing on endpoint and quantitative RT‐PCR using mRNA. The advantages and limitations of these along with alternative methods will be discussed, and how they may be further developed to advance forensic body fluid identification. This article is categorized under: Forensic Biology > Body Fluid Identification Forensic Biology > Applications of RNA