基于ARMS多重荧光复合扩增技术的人源肿瘤细胞系鉴定体系研究

人源肿瘤细胞系是医学生物学各领域使用的最重要生物资源之一。但我国细胞系交叉污染问题极为严重，威胁着获取的研究数据和结论的有效性。本课题拟以ARMS-PCR方法结合多重荧光复合扩增技术为基础，建立同步鉴定具有高度多态性的STR和双等位基因特异分型的SNP两种遗传标记的检测体系，以准确鉴别肿瘤细胞系身份真实性，解决依赖于STR分型的细胞身份检测中，因高频微卫星不稳定、杂合性丢失、AMEL基因座缺失或突变导致的Y染色体信号丢失及不均衡混合等造成的检测局限。并在较大规模进行该检测体系的灵敏度、准确性、特异性等性能验证及结果验证和应用性验证，从方法学上建立结果准确、操作简便、易于推广使用的肿瘤细胞系身份新型检测体系，为肿瘤细胞系这一重要科研资源提供可靠质量保障。并以此为基础系统性研究国内肿瘤细胞系身份真实性及交叉污染情况。建立肿瘤细胞系身份鉴定检测数据库，为肿瘤细胞系使用提供更为全面的比对依据。

Human tumor cell line is one of the most important biological resources in diverse fields of biomedical biology. However, the cross-contamination problem is critical in the research fields of our country, which threatens the accuracy of the obtained data. In this study, synchronous detection system will be established for two genetic markers, including STR with high polymorphism and SNP with double allele specific typing, based on fluorescent multiplex ARMS technology, in order to break through the detection limitation of STR technology caused by MSI-H, Y chromosome signal loss caused by AMEL locus deletion or mutation, unstable heterozygosity loss, and unbalanced mixed samples. In addition, the performance verification of the detection system, including sensitivity, accuracy and specificity, and its application will be evaluated on a large scale. From the perspective of methodology, this novel identification system has simple operation procedures while obtaining accurate conclusion at the same time, and provide reliable quality assurance for the usage of tumor cell lines as an important scientific research resource. The novel detection system has extensive applications in the scientific research and will provide comprehensive and accurate evaluation of the identification of tumor cell lines used domestically and exploration of the source of cell line cross-contamination. The identification database of tumor cell lines will be established based on the application of the novel detection system, to provide a more comprehensive comparison basis for the usage of human tumor cell lines in the scientific research community.