Characterisation of body fluid specific microRNA markers by capillary electrophoresis

Abstract

The characterisation of RNA molecules for the purpose of body fluid identification is currently a major field in forensic genetics; with a great deal of effort going towards the analysis of messenger RNA (mRNA). There is also some effort with targeting microRNA (miRNA) which is a more stable RNA molecule than mRNA; due to its short size and role in RNA interference. Most research into forensic miRNA analysis is based around quantitative PCR (qPCR). No substantial research has yet been carried out on capillary electrophoretic (CE) analysis of miRNA. Thus the aim of this study was to explore the viability of CE of miRNA. Samples of blood, saliva, semen, and vaginal material were obtained from a number of volunteers with their informed consent. All samples then underwent standard DNA extraction using QIAAmp DNA mini kit. cDNA synthesis was carried out using stem-loop reverse transcription and commercially available stem-loop primers. qPCR was performed using a 7500 Fast Real-Time PCR Machine and commercially available miRNA assays. The amplified product then underwent fragment analysis using an ABI 3130 genetic analyser. The findings have demonstrated that CE analysis of miRNA markers could be viable for the purpose of forensic genetics. The fragment sizes (between 30 and 70bp) suggest that such CE based miRNA assays could be multiplexed with STR kits with minimal modification; thus enhancing the capability of DNA profiling.