高通量测序样本准备方法及其在孕妇外周血微量循环核酸研究中的应用

项目名称： 高通量测序样本准备方法及其在孕妇外周血微量循环核酸研究中的应用

项目编号： No.61271055

项目类型： 面上项目

立项/批准年度： 2013

项目学科： 无线电电子学、电信技术

项目作者： 葛芹玉

作者单位： 东南大学

项目金额： 80万元

中文摘要： 近年来循环核酸在肿瘤、产前筛查等领域得到了极大的关注。利用高通量测序手段在全基因组水平上对循环核酸进行系统的研究分析对于其在疾病的早期诊断、分期、监测、预后判断等方面更好的应用意义重大。本项目结合高通量测序技术平台提出了一种低样本输入量、快速、高效的集文库制备与模板扩增为一体的测序样本准备方案，同时优化改良血浆中循环核酸的提取方法，结合生物信息分析平台，对不同妊娠时期和几种异常妊娠状态下孕妇外周血浆中cMVs（Circulating Microvesicles）内外的循环DNA/RNA/miRNA进行全基因组范围的差异表达分析。用以阐述循环DNA的大小、分布及存在规律等基础生物学特性，并试图对miRNA与mRNA/DNA之间的相互调控关系及其与妊娠等的关联进行更加全面的解释，为循环核酸用于无创性产前筛查与诊断提供新的思路和研究平台。本课题研究在功能基因组学，分子遗传学及生物医学领域。

中文关键词： 高通量测序；循环核酸；文库制备；孕妇；肺癌

英文摘要： Circulating nucleic acid are getting a lot of attentions in cancer research and prenatal diagnosis. It must be more significant to detect circulating nucleic acid using the next generation sequencing technology which could made its application in early diagnosis, staging, monitoring and prognosis more comprehensive and reliable. In this present proposal, we propose a low input, quick and effective next generation sequencing sample preparation method which set library preparartion and sequencing template amplification in one on the surface of micro-beads. The circulating nucleic acid extraction method was also modified and improved. Circulating DNA/RNA/miRNA in and out of the cMVs (Circulating Microvesicles) from pregnant women in their different gestation stages and abnormal status were sequenced and analyzed. We attempt to explain the basic characteristics such as fragment sizes, distribution and existing rules of circulating DNA using bioinformatics method based on these sequencing data, as well as the whole regulating relations between miRNA and RNA or DNA under certain pregnancy. This provides new way and research platform for circulating nucleic acid and non-invasive prenatal diagnosis. The project has wide application in founctional genomics, molecular geneitics and biomedical science.

英文关键词： NGS；Circulating Nucleic Acid；Library Preparation；Pregnancy；Lung Cancer