微量接触类、混合法医生物物证快速检验策略研究

Recently, the amount of trace / touch DNA and biological mixtures have increased greatly, and have been the subjects of much research. It is difficult to retrieve usable profiles from such samples and repeated experiments are needed, which may slow down the whole process. Skin or mucosa cells can be left on items of the crime scene which are called trace / touch DNA evidence. Such as knife handle, key, socks, toothbrush and so on. It is difficult to see the location of cells or DNA, such evidence usually be processed blindly. If too much samples are cut, DNA concentration could be decreased, sometimes contamination problem may arise. Biological mixtures are composed of samples from different donor which usually appear on such cases like rape and murder. Mixed DNA profile may be recovered by routine method. It is difficult to identify a suspect or search matches in the DNA database using a mixed DNA profile. .In this study, we aim to develop a strategy of high speed sample disposal and DNA amplification based on previous research achievements. Systematic study will be carried out first to investigate a series of basic data and theory related to casework biological samples which could assist the sample disposal of given crime scene scenarios. For example, how much DNA may deposits on a wide range of substrates and variation of storage time. To explore the source of the touch DNA and the occurrence of secondary transfer of DNA, to investigate cell morphology and storage time on different substrates. Then, procedure for rapid PCR will be developed that suited to the commonly used forensic STR kits, including the selection of rapid DNA polymerase and additives, rapid amplification procedure, and validation of the system. The basic data investigation will be combined with techniques such as, rapid PCR method to develop a strategy of high speed sample disposal and DNA amplification. The strategy can assist police investigations by rapidly deriving DNA information from touch DNA and biological mixture sample left by perpetrators. The value of biological evidence can be better utilized.

微量接触类、混合生物检材是目前检案中经常遇到的物证,由于检验成功率低，往往需要反复多次检验，成为限速步骤。微量接触类物证是皮肤或粘膜与载体接触而形成，如刀柄、钥匙、手机、鞋袜、手套、牙刷等，检验时很难判断载体上样本的具体部位，往往盲目剪取，剪取面积过大时又会降低DNA浓度或引入污染。混合生物物证是指由不同供者的生物样本组成的物证，常见于强奸、杀人等案件，常规方法往往得到混合DNA图谱，既不能认定嫌疑人，也难入DNA数据库比对查找。本项目拟在前期基础上，从物证的快速处置和扩增两个环节入手，通过系统探究生物物证中的DNA含量、保存时间、来源、二次转移、常见细胞的特征、识别等基础理论和数据问题，同时从聚合酶、快速PCR扩增仪、添加剂等入手，研究建立适合法医常用复合STR体系的半小时快速扩增技术。最终形成适合此类物证的快速处置和检验策略，从而提高法医检验的效率，提升生物物证的证据价值和证明力。

[背景]微量接触类、混合生物检材是常见的现场物证，检验成功率低，且需反复检验，成为限速步骤。接触类物证是皮肤或粘膜与载体接触而形成，如刀柄、钥匙等，检验时很难判断样本的精确部位，盲目剪取容易降低DNA浓度或引入污染。混合生物物证是指由不同供者的生物样本组成的物证，常见于强奸、杀人等案件，常规方法往往得到混合DNA图谱，难以认定嫌疑人。急需开展基础理论研究，建立快速处置方法。[主要内容]1.微量接触类检材快速处理策略研究。研究渗透性、非渗透性等不同载体上接触类DNA的含量研究；DNA来源和二次转移问题；多种物证检验时最佳方案。2.混合生物检材快速处理策略研究。包括：不同供者DNA的来源研究；常见细胞的形态特性研究、标记识别和分离技术研究；微量细胞DNA的提取和扩增一体化技术研究。3.快速扩增技术研究。快速扩增DNA聚合酶的选择、添加剂的选择、快速扩增体系构建和程序的优化与验证等研究。[结果和数据]1.渗透性载体的DNA检测量(100%)高于非渗透性载体的DNA检测量(90%)。2.渗透性载体物证用直接剪取和真空吸附的效果较好，非渗透性载体物证用胶带粘取和两步擦拭效果较好。3.接触类物证常规保存，在360天以内，DNA检出量无明显差异。4.物证中的DNA转移到物证袋的发生率为100%，且在纸质（26.27%）和塑料物证袋（17.85%）中的转移率有差别。5.微量接触类检材的DNA来源于游离DNA和细胞内DNA，其中游离DNA约占总DNA量的12.9%。6.建立了针对口腔上皮细胞和精子细胞的染色标记方法。不同载体上的细胞随着保存时间的延长细胞形态未见明显变化，但DNA检出率呈下降趋势。7.针对混合样本检验，建立了咬痕、男女混合样本、轮奸案中混合精斑的分离检验方法，建立了适合微量细胞的裂解、扩增一体化1微升体系。8.建立了基于Identifiler Plus试剂、Yfiler试剂引物、DNA Typer15 plus试剂的10μL扩增体系和快速扩增程序，耗时在1小时左右。9.针对本研究涉及的物证，编制了快速处置和检验策略。[科学意义]系统探究了法医常见生物物证中的细胞和DNA的存在状态、检出量等基础数据和理论问题，为法医生物物证的处置和检验提供科学指导。并以此为基础，从物证的提取、处置、扩增检验等环节展开系统研究，建立了系统的快速处置和检验策略,提升了证据价值。