基于DNA甲基化的三种常见体液组织来源鉴定

Source identification of three types of body fluids based on DNA methylation

目的针对唾液、精液、外周血,找到一组可用于区分不同体液的组织差异性甲基化位点,为确定案件现场提取的体液组织来源提供科学依据。方法选取常见体液样本共49份,运用Illumina 850K甲基化芯片进行全基因组甲基化检测,筛选获得17个候选CpG位点;使用焦磷酸测序对55份样本进行测序,实际共检测了候选位点及其附近的位点共34个CpG位点。选择基于赤池信息量(AIC)的逐步条件多分类逻辑回归方法进行数据分析。结果基于3个CpG位点(cg25373595、cg18121066、cg17283169)构建了多分类逻辑回归模型用于体液组织来源预测,分别位于RAP1GAP2,TBCD,CALML3基因上。该模型对精液来源预测的AUC、灵敏度和特异性均为1,对唾液和静脉血来源的预测AUC和灵敏度均为1,特异性分别为0.99和0.98。在独立的唾液、精液和静脉血3种体液共24份样本中对该预测模型进行验证,预测结果全部正确。结论本文建立的基于3个CpG位点的体液组织来源鉴定方法可以实现唾液、精液和静脉血3种体液的有效区分,在法庭科学中具有潜在的应用价值。

Objective To identify a group of tissue differential methylation sites which can be used to distinguish different types of body fluids, such as saliva, semen and venous blood, and provide scientific basis for determining the source of body fluids extracted from the case scene. Methods Forty-nine different samples(including venous blood, saliva,semen, menstrual blood and vaginal secretion) were tested using Infinium Human Methylation EPIC BeadArray(850 K)methylation chip, and 17 CpG sites were initially screened as tissue-specific sites. Pyrosequencing was used to verify these 17 sites and their adjacent sites in 55 samples. For data analysis, stepwise multinomial logistic regression analysis was carried out based on Akaike Information(AIC). Results We identified 3 novel tissue-specific methylation sites: cg25373595,cg18121066, and cg17283169, which were located on RAP1 GAP2, TBCD, and CALML3 genes respectively. A multinomial logistic regression model based on the three-CpG sites was constructed to predict the source of different body fluid spots. The area under the curve(AUC), sensitivity and specificity of the model for the prediction of semen were 1. The AUC and sensitivity for predicting saliva and blood were 1, and the specificity was 0.99 and 0.98 respectively. The performance of 3-CpG model was further validated in 24 independent samples, and all prediction results were accurate. Conclusion The 3-CpG model can effectively distinguish saliva, semen and venous blood, which has potential applications in forensic science.