拷贝数变异在中国遗传性耳聋人群中的分布及筛查策略研究

项目名称： 拷贝数变异在中国遗传性耳聋人群中的分布及筛查策略研究

项目编号： No.81500801

项目类型： 青年科学基金项目

立项/批准年度： 2016

项目学科： 医药、卫生

项目作者： 郭荦

作者单位： 复旦大学

项目金额： 18万元

中文摘要： 非综合症型耳聋（NSHL）具有很高的遗传异质性，因此基因组靶向捕获和高通量测序已成为耳聋突变筛查的有效方法。然而，以往的研究在分析高通量测序数据时大多只侧重于发现点突变和小片段插入缺失，却忽略了耳聋基因的拷贝数变异（CNV）。CNV与遗传性聋密切相关，然而限于分辨率和成本等因素的制约，经典方法不适用于耳聋基因CNV的筛查。本项目拟应用多重PCR扩增结合高通量测序的方法（Ampliseq），实现耳聋基因点突变和CNV的同步筛查。我们将使用Ampliseq产生的覆盖倍数数据进行CNV calling，采用实时定量PCR、断裂位点定位验证Ampliseq检测CNV的可靠性；并结合生物信息学方法分析耳聋基因序列，预测CNV热点区域，对靶向捕获方案进行优化，提高Ampliseq检测CNV的准确度和效率。本研究将为建立更全面的遗传性聋突变筛查策略提供实验基础。

中文关键词： 致病基因；基因突变；拷贝数变异；突变筛查；高通量测序

英文摘要： The genetic heterogeneity of non-syndrome hearing loss (NSHL) is very high, therefore target genome enrichment coupled with high throughput sequencing has been implemented for mutation screening of NSHL. However, previous studies using high throughput sequencing strategies focused on identifying point mutations and small INDELs, while neglecting the gene copy number variations (CNV). CNV could cause hereditary hearing loss, however, due to the limited resolution or the high price, the classic CNV screening methods is not suitable for identifying CNVs in hearing loss genes. This project intends to screening for small mutations and CNVs simultaneously using ultra multiplex PCR and high through put sequencing technics (Ampliseq). We will making CNV callings using the depth of coverage data generated by Ampliseq, then verify the CNV callings using real-time quantitative PCR, and breakpoint mappings. The primers used in Ampliseq will be optimized by sequence architecture analysis of hearing loss genes. This study will provide experimental basis for establishing a more efficient and more comprehensive mutation screening strategy for hereditary hearing loss.

英文关键词： pathogenic genes;gene mutations;copy number variants;mutation screening;massively parallel sequencing