The accurate characterization of triplet repeats, especially the overrepresented CAG repeats, is increasingly relevant for several reasons. First, germline expansion of CAG repeats above a gene-specific threshold causes multiple neurodegenerative disorders; for instance, Huntington's disease (HD) is triggered by >36 CAG repeats in the huntingtin (HTT) gene. Second, extreme expansions up to 800 CAG repeats have been found in specific cell types affected by the disease. Third, synonymous single nucleotide variants within the CAG repeat stretch influence the age of disease onset. Thus, new sequencing-based protocols that profile both the length and the exact nucleotide sequence of triplet repeats are crucial. Various strategies to enrich the target gene over the background, along with sequencing platforms and bioinformatic pipelines, are under development. This review discusses the concepts, challenges, and methodological opportunities for analyzing triplet repeats, using HD as a case study. Starting with traditional approaches, we will explore how sequencing-based methods have evolved to meet increasing scientific demands. We will also highlight experimental and bioinformatic challenges, aiming to provide a guide for accurate triplet repeat characterization for diagnostic and therapeutic purposes.

中文翻译：

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驾驭三联体重复测序：亨廷顿病的概念、方法学挑战和前景。


由于多种原因，三重体重复序列的准确表征，尤其是过度代表的 CAG 重复序列，越来越重要。首先，CAG 重复序列的种系扩增超过基因特异性阈值会导致多种神经退行性疾病;例如，亨廷顿病 （HD） 是由亨廷顿蛋白 （HTT） 基因中的 >36 CAG 重复序列触发的。其次，在受疾病影响的特定细胞类型中发现了高达 800 个 CAG 重复序列的极端扩增。第三，CAG 重复序列中的同义单核苷酸变异会影响发病年龄。因此，分析三联体重复序列的长度和确切核苷酸序列的基于测序的新方案至关重要。在背景上富集靶基因的各种策略，以及测序平台和生物信息学管道，正在开发中。本文以 HD 为案例研究，讨论了分析三重体重复的概念、挑战和方法学机会。从传统方法开始，我们将探索基于测序的方法如何发展以满足日益增长的科学需求。我们还将重点介绍实验和生物信息学挑战，旨在为诊断和治疗目的的准确三联体重复表征提供指南。