Tekin Demet, Yan Denise, Bademci Guney, Feng Yong, Guo Shengru, Foster Joseph, Blanton Susan, Tekin Mustafa, Liu Xuezhong
Department of Otolaryngology, University of Miami Miller School of Medicine, Miami, FL 33136, USA.
Dr. John T. Macdonald Foundation Department of Human Genetics, and John P. Hussman Institute for Human Genomics, University of Miami, Miami, FL 33136, USA.
Hear Res. 2016 Mar;333:179-184. doi: 10.1016/j.heares.2016.01.018. Epub 2016 Feb 2.
Extreme genetic heterogeneity along with remarkable variation in the distribution of causative variants across in different ethnicities makes single gene testing inefficient for hearing loss. We developed a custom capture/next-generation sequencing gene panel of 146 known deafness genes with a total target size of approximately 1 MB. The genes were identified by searching databases including Hereditary Hearing Loss Homepage, the Human Genome Mutation Database (HGMD), Online Mendelian Inheritance in Man (OMIM) and most recent peer-reviewed publications related to the genetics of deafness. The design covered all coding exons, UTRs and 25 bases of intronic flanking sequences for each exon. To validate our panel, we used 6 positive controls with variants in known deafness genes and 8 unsolved samples from individuals with hearing loss. Mean coverage of the targeted exons was 697X. On average, each sample had 99.8%, 96.2% and 92.7% of the targeted region coverage of 1X, 50X and 100X reads, respectively. Analysis detected all known variants in nuclear genes. These results prove the accuracy and reliability of the custom capture experiment.
极高的基因异质性，以及不同种族中致病变异分布的显著差异，使得单基因检测对于听力损失而言效率低下。我们开发了一个包含146个已知耳聋基因的定制捕获/新一代测序基因panel，总目标大小约为1MB。这些基因是通过搜索包括遗传性听力损失主页、人类基因组突变数据库（HGMD）、《人类孟德尔遗传》（OMIM）以及与耳聋遗传学相关的最新同行评审出版物在内的数据库来确定的。该设计涵盖了每个外显子的所有编码外显子、非翻译区（UTR）以及内含子侧翼序列的25个碱基。为了验证我们的panel，我们使用了6个在已知耳聋基因中有变异的阳性对照以及8个来自听力损失个体的未解决样本。目标外显子的平均覆盖度为697倍。平均而言，每个样本在1倍、50倍和100倍读数下分别有99.8%、96.2%和92.7%的目标区域覆盖度。分析检测到了核基因中的所有已知变异。这些结果证明了定制捕获实验的准确性和可靠性。
