Characterisation of OFMIR-205,MIR-451 and RNU44 in blood and saliva samples

Over the past 20 years, RNA analysis has played an important role in forensic body fluid identification (BFID). MicroRNA (miRNA, miR) has begun to take the forefront over messenger RNA BFID as its theoretical stability makes it more suited to forensic samples. 1-5 Current research in this area have identified non-coding RNA and small nucleolar RNA (snoRNA) for the resolution and accurate normalisation of body fluids.2-7 Here we explore the sensitivity, specificity and applications of miR-205, miR-451 and RNU44 using single and mixed body fluids. An in-house miRNA protocol involving DNA extraction, cDNA synthesis (TaqMan probes) and qPCR analysis (relative quantification) was performed on blood and saliva samples using body fluid specific probes miR-205 (saliva), miR-451 (blood) and reference gene RNU44.2,6, All samples were analysed and presented following the MIQE guidelines for qPCR experiments.8 Blood and saliva mixtures were resolved using miR-205, miR-451 and RNU44. Limit of detection studies indicate a greater sensitivity with the body fluid specific miRNA than with the reference gene. Species specificity was seen using the reference gene but not with the body fluid- specific probes. References: [1] R. Fleming, S. Harbison (2010) FSI: Genetics 4(4): 244-256. [2] D. Zubakov et al. (2010) Int J Legal Med 124(3): 217-226. [3] Hanson et al. (2009) Anal Biochem 387(2): 303-314. [4] E. Sauer, B. Madea, C. Courts (2014) FSI: Genetics (Accepted manuscript). [5] M. L. Uchimoto, et al. (2013) FSI: Genetics 7(4): 418-21. [6] E. J. Omelia, M.L. Uchimoto, G. Williams (2013) Anal. Biochem. 435(2):120-122. [7] Life Technologies Application note: TaqMan microRNA assays. [8] S.A. Bustin et al. (2009) Clin.Chem. 55(4): 611-622.