Precise estimation of postmortem interval (PMI) is crucial in some criminal cases. This study aims to find some optimal markers for PMI estimation and build a mathematical model that could be used in various temperature conditions. Different mRNA and microRNA markers in rat brain samples were detected using real-time fluorescent quantitative PCR at 12 time points within 144 h postmortem and at temperatures of 4, 15, 25, and 35 °C. Samples from 36 other rats were used to verify the animal mathematical model. Brain-specific mir-9 and mir-125b are effective endogenous control markers that are not affected by PMI up to 144 h postmortem under these temperatures, whereas the commonly used U6 is not a suitable endogenous control in this study. Among all the candidate markers, ΔCt (β-actin) has the best correlation coefficient with PMI and was used to build a new model using R software which can simultaneously manage both PMI and temperature parameters. This animal mathematical model is verified using samples from 36 other rats and shows increased accuracy for higher temperatures and longer PMI. In this study, β-actin was found to be an optimal marker to estimate PMI and some other markers were found to be suitable to act as endogenous controls. Additionally, we have used R code software to build a model of PMI estimation that could be used in various temperature conditions.
This is a preview of subscription content, access via your institution.
Buy single article
Instant access to the full article PDF.
Tax calculation will be finalised during checkout.
Henssge C, Madea B. Estimation of the time since death in the early post-mortem period. Forensic Sci Int. 2004;144:167–75.
Thyssen PJ, de Souza CM, Shimamoto PM, Salewski TDB, Moretti TC. Rates of development of immatures of three species of Chrysomya (Diptera: Calliphoridae) reared in different types of animal tissues: implications for estimating the postmortem interval. Parasitol Res. 2014;113:3373–80.
Kikuchi K, Kawahara KI, Biswas KK, Ito T, Tancharoen S, Shiomi N, et al. HMGB1: a new marker for estimation of the postmortem interval. Exp Ther Med. 2010;1:109–11.
Dorandeu A, Lorin DLGG. Contribution of the TUNEL method for post-mortem interval estimation: an experimental study. Ann Pathol. 2013;33:80–3.
Sampaio-Silva F, Magalhaes T, Carvalho F, Dinis-Oliveira RJ, Silvestre R. Profiling of RNA degradation for estimation of post morterm interval. PLoS One. 2013;8:e56507.
Li WC, Ma KJ, Lv YH, Zhang P, Pan H, Zhang H, et al. Postmortem interval determination using 18S-rRNA and microRNA. Sci Justice. 2014;54:307–10.
Lv YH, Ma KJ, Zhang H, He M, Zhang P, Shen YW, et al. A time course study demonstrating mRNA, microRNA, 18S rRNA, and U6 snRNA changes to estimate PMI in deceased rat’s spleen. J Forensic Sci. 2014;59:1286–94.
Chen PS, Su JL, Cha ST, Tarn WY, Wang MY, Hsu HC, et al. miR-107 promotes tumor progression by targeting the let-7 microRNA in mice and humans. J Clin Invest. 2011;121:3442–55.
Lardizabal MN, Nocito AL, Daniele SM, Ornella LA, Palatnik JF, Veggi LM. Reference genes for real-time PCR quantification of microRNAs and messenger RNAs in rat models of hepatotoxicity. PLoS One. 2012;7:e36323.
Tea M, Michael MZ, Brereton HM, Williams KA. Stability of small non-coding RNA reference gene expression in the rat retina during exposure to cyclic hyperoxia. Mol Vis. 2013;19:501–8.
Griffiths-Jones S, Grocock RJ, van Dongen S, Bateman A, Enright AJ. miRBase: microRNA sequences, targets and gene nomenclature. Nucl Acids Res. 2006;34:D140–4.
Ecsedi M, Rausch M, Grosshans H. The let-7 microRNA directs vulval development through a single target. Dev Cell. 2015;32:335–44.
Bustin SA, Benes V, Garson JA, Hellemans J, Huggett J, Kubista M, et al. The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments. Clin Chem. 2009;55:611–22.
Birdsill AC, Walker DG, Lue L, Sue LI, Beach TG. Postmortem interval effect on RNA and gene expression in human brain tissue. Cell Tissue Bank. 2011;12:311–8.
Zhang H, Zhang P, Ma KJ, Lv YH, Li WC, Luo CL, et al. The selection of endogenous genes in human postmortem tissues. Sci Justice. 2013;53:115–20.
Smart JL, Kaliszan M. Use of a finite element model of heat transport in the human eye to predict time of death. J Forensic Sci. 2013;58(Suppl 1):S69–77.
Muggenthaler H, Sinicina I, Hubig M, Mall G. Database of post-mortem rectal cooling cases under strictly controlled conditions: a useful tool in death time estimation. Int J Legal Med. 2012;126:79–87.
Vennemann M, Koppelkamm A. mRNA profiling in forensic genetics I: possibilities and limitations. Forensic Sci Int. 2010;203:71–5.
Odriozola A, Riancho JA, de la Vega R, Agudo G, Garcia-Blanco A, de Cos E, et al. miRNA analysis in vitreous humor to determine the time of death: a proof-of-concept pilot study. Int J Legal Med. 2013;127:573–8.
Zapico CS, Menendez ST, Nunez P. Cell death proteins as markers of early postmortem interval. Cell Mol Life Sci. 2014;71:2957–62.
Inoue H, Kimura A, Tuji T. Degradation profile of mRNA in a dead rat body: basic semi-quantification study. Forensic Sci Int. 2002;130:127–32.
Gonzalez-Herrera L, Valenzuela A, Marchal JA, Lorente JA, Villanueva E. Studies on RNA integrity and gene expression in human myocardial tissue, pericardial fluid and blood, and its postmortem stability. Forensic Sci Int. 2013;232:218–28.
Young ST, Wells JD, Hobbs GR, Bishop CP. Estimating postmortem interval using RNA degradation and morphological changes in tooth pulp. Forensic Sci Int. 2013;229:161–3.
Vennemann M, Koppelkamm A. Postmortem mRNA profiling II: practical considerations. Forensic Sci Int. 2010;203:76–82.
Mocellin S, Rossi CR, Pilati P, Nitti D, Marincola FM. Quantitative real-time PCR: a powerful ally in cancer research. Trends Mol Med. 2003;9:189–95.
Neville MJ, Collins JM, Gloyn AL, McCarthy MI, Karpe F. Comprehensive human adipose tissue mRNA and microRNA endogenous control selection for quantitative real-time-PCR normalization. Obesity (Silver Spring). 2011;19:888–92.
Burke JE, Sashital DG, Zuo X, Wang YX, Butcher SE. Structure of the yeast U2/U6 snRNA complex. RNA. 2012;18:673–83.
Benz F, Roderburg C, Vargas CD, Vucur M, Gautheron J, Koch A, et al. U6 is unsuitable for normalization of serum miRNA levels in patients with sepsis or liver fibrosis. Exp Mol Med. 2013;45:e42.
Xiang M, Zeng Y, Yang R, Xu H, Chen Z, Zhong J, et al. U6 is not a suitable endogenous control for the quantification of circulating microRNAs. Biochem Biophys Res Commun. 2014;454:210–4.
Munoz-Barus JI, Rodriguez-Calvo MS, Suarez-Penaranda JM, Vieira DN, Cadarso-Suarez C, Febrero-Bande M. PMICALC: an R code-based software for estimating post-mortem interval (PMI) compatible with Windows, Mac and Linux operating systems. Forensic Sci Int. 2010;194:49–52.
About this article
Cite this article
Ma, J., Pan, H., Zeng, Y. et al. Exploration of the R code-based mathematical model for PMI estimation using profiling of RNA degradation in rat brain tissue at different temperatures. Forensic Sci Med Pathol 11, 530–537 (2015). https://doi.org/10.1007/s12024-015-9703-7
- Postmortem interval (PMI)
- Reverse transcription quantitative real-time PCR
- R code