Evaluation of RNA Stability and Molecular Biomarkers for Post-Mortem Interval Estimation in Rat Organs

Post-mortem interval (PMI) analysis plays a crucial role in forensic investigations, providing essential insights into the time since death. This study examined histological changes and ribonucleic acid (RNA) quantification across major organs to identify molecular indicators for PMI estimation. Because RNA gradually degrades after death, understanding its stability under different tissue, temperature, and PMI conditions provides valuable forensic insights. We analyzed post-mortem changes in total RNA from the heart, kidney, liver, and lung tissues of Sprague Dawley rats stored at 4 °C and 26 °C. Tissue samples were collected at various PMIs and evaluated histologically for cellular integrity. Total RNA concentration and purity were measured, and complementary DNA (cDNA) was synthesized for molecular analysis. Expression levels of 5S rRNA, <i>B2m</i>, <i>Gapdh</i>, and <i>Sort1</i> were quantified using reverse transcription quantitative PCR (RT-qPCR). The results showed that PMI and organ type significantly affected total RNA concentration, whereas temperature exerted only a minor effect. Among the four target genes, 5S rRNA exhibited the lowest Ct values, indicating the highest stability. Notably, RNA degradation patterns varied with temperature, particularly in kidney and liver tissues. These findings suggest that RNA-based molecular markers, particularly 5S rRNA, may serve as promising indicators for accurate PMI estimation.