Cell free DNA and MiRNA analysis by quantitative Real-Time polymerase chain reaction in postmortem interval determination.

The time elapsed after death is known as the postmortem interval (PMI). After death, the body undergoes changes such as algor mortis, rigor mortis, livor mortis, and putrefaction. In forensic medicine, PMI is traditionally determined using standard methods and forensic entomology, both of which can be affected by environmental conditions. Recently, genetic materials, which are less influenced by the environment, have gained attention in PMI studies. Cell-free DNA (cfDNA) comprises short DNA fragments released into the bloodstream due to cell necrosis and apoptosis. This study quantitatively analyzed cfDNA and specific microRNAs (miRNAs) - miR-122, miR-133, miR-1, miR-16, and let-7e - in serum samples from rats at various time intervals to assess their correlation with PMI. The results were analyzed using Ct, ΔCt, ΔΔCt, and fold change methods. Statistically, miR-16 and miR-122 had lower ΔCt values at all time points. miR-16 showed a lower ΔΔCt average at 24 and 48 h and the highest fold change at 24 h. Reference genes GAPDH and ACTB declined over time, with higher averages at 0, 6, and 12 h compared to 24 and 48 h. This research is the first to quantitatively explore cfDNA's potential role in PMI estimation.