小鼠断颈死后组织中RNA与DNA相对含量与死亡时间相关性的继续研究

Estmation the early postmortem interval (PMI) is very diffcult in forensic appliction and study. Previous research has drawn confilcting conclusions about the postmortem stability of RNA in brain tissue. Many studies report that global measures of total RNA quality are relative stable over a wide range of PMI while some others report a definite loss with increasing time. And the studies of subtypes of RNA have also had a varying conclusions, with some indicating generalized stability while others have reported significant loss of a set of individual mRNA transcripts with increasing PMI. Our previous project has studied the change of mRNA under the control of DNA in the mouse brain tissue and heart tissue within 48 hours of PMI in 37℃. A significant loss of mRNA transcripts in the range of 2-24 hours, but we could not intepret it with a better regression curve because the PMI points in this range is not enough. More PMI points in the range of 2-24 hours and more target mRNA need to be studied in the next project to prove the change of mRNA in the early PMI. We plan to measure the change of 22 mRNA in the 24 hours of PMI under the control of DNA in 37℃ and 18℃ (average tempreature in Chengdu) to determine how PMI affects the expression of individual genes and in order to determine whether there is a line relationship between PMI and mRNA. In our previous project we also find that microRNAs are very stable in 48 hours of PMI, some microRNA will begin to loss at 48 hours of PMI. We will study the change of microRNA under the controls of DNA in 240 hours of PMI in outdoor winter and summer of Chengdu in the next project. A detailed story about changes of mRNA in 24 hours and microRNA in 240 hours of PMI will be wrote after we finished next project. And the result will be tested by brain and heart tissues from at least 100 autopsy with different PMI.Better regression curves will be drawn by the intimate datas, and the could be used to estimate the PMI. The result of this study can also help other scientist to study the RNA integrity in postmortem and further study in the estimation of PMI.

死亡时间推断是法医学面临的难题之一。RNA在机体死后早期是否具有规律性变化，能否用于推断死亡时间，在学术界仍有争议。课题组前期以DNA为参照研究RNA在死后48小时内含量变化，证明37℃环境下机体死亡2-24小时内RNA含量有明显时间相关降解，并在2013年国际法医遗传学会议中进行大会报告。前期研究在该范围内设置时间点的密度和目的基因数量不足，无法获得准确的线性关系。课题组计划继续以DNA为参照在前期研究基础上对机体死亡后37℃和18℃环境中24小时内高密度时间点分组的22个目的基因，以及在成都冬夏两季室外环境机体死亡后240小时内5个microRNA含量变化进行深入研究。以期阐明RNA/DNA相对含量在机体死后24小时内详细的变化规律，在上述争议中提出有力证据。继而进行法医学尸体解剖样本研究，对本研究所获得规律性结论的实践应用价值进行评估，为法医学死亡时间推断提供解决思路和理论基础。

死亡时间（Postmortem Interval, PMI）推断一直是法医学实践的重要内容，是法医学研究的重点方向。本研究通过检测死后小鼠心脏以及大脑组织中基因组DNA以及mRNA表达量，发现基因组DNA的CT值与0-48h的死亡时间没有明显的相关性，作为内参时， mRNA的表达量随死亡时间延长而降低，呈现出与死亡时间显著的相关性，并确定了表达量与死亡时间的回归曲线。其次，死后脑、心组织中mRNA降解速度具有时间阶段性，总体上，死亡早期mRNA降解速度快于中期。第三，同一组织中的相同mRNA基因，在片段大小为79bp-365bp时，其在0-48h PMI的降解速度与片段大小无统计学差异。第四，mRNA的降解具有组织差异性，加上对国内外研究动态的了解，课题组认为联合研究多基因在死后多种组织中的变化情况，更有利于准确判断死亡时间。