1Biomedical Informatics, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio 45229, USA.
BMC Biotechnol. 2010 Feb 10;10:10. doi: 10.1186/1472-6750-10-10.
Despite current knowledge of mutations in 45 genes that can cause nonsyndromic sensorineural hearing loss (SNHL), no unified clinical test has been developed that can comprehensively detect mutations in multiple genes. We therefore designed Affymetrix resequencing microarrays capable of resequencing 13 genes mutated in SNHL (GJB2, GJB6, CDH23, KCNE1, KCNQ1, MYO7A, OTOF, PDS, MYO6, SLC26A5, TMIE, TMPRSS3, USH1C). We present results from hearing loss arrays developed in two different research facilities and highlight some of the approaches we adopted to enhance the applicability of resequencing arrays in a clinical setting.
We leveraged sequence and intensity pattern features responsible for diminished coverage and accuracy and developed a novel algorithm, sPROFILER, which resolved >80% of no-calls from GSEQ and allowed 99.6% (range: 99.2-99.8%) of sequence to be called, while maintaining overall accuracy at >99.8% based upon dideoxy sequencing comparison.
Together, these findings provide insight into critical issues for disease-centered resequencing protocols suitable for clinical application and support the use of array-based resequencing technology as a valuable molecular diagnostic tool for pediatric SNHL and other genetic diseases with substantial genetic heterogeneity.
尽管目前已经了解了 45 个可以导致非综合征性感音神经性听力损失（SNHL）的基因突变，但尚未开发出一种能够全面检测多个基因突变的统一临床检测方法。因此，我们设计了 Affymetrix 重测序微阵列，可以对 13 个发生 SNHL 突变的基因（GJB2、GJB6、CDH23、KCNE1、KCNQ1、MYO7A、OTOF、PDS、MYO6、SLC26A5、TMIE、TMPRSS3、USH1C）进行重测序。我们展示了在两个不同研究机构开发的听力损失阵列的结果，并强调了我们采用的一些方法，以增强重测序阵列在临床环境中的适用性。
我们利用负责降低覆盖度和准确性的序列和强度模式特征，开发了一种新算法 sPROFILER，该算法解决了 GSEQ 中超过 80%的无呼叫问题，并允许 99.6%（范围：99.2-99.8%）的序列被调用，同时保持整体准确性>99.8%，基于双脱氧测序比较。
这些发现共同为适合临床应用的以疾病为中心的重测序方案提供了重要问题的深入了解，并支持使用基于阵列的重测序技术作为小儿 SNHL 和其他具有大量遗传异质性的遗传疾病的有价值的分子诊断工具。
