Background Presence of excess unaltered, wild-type DNA (wtDNA) providing information of little clinical value may often mask low-level mutations containing important diagnostic or therapeutic clues. This is a recurring hurdle in biotechnology and medicine, including cancer, prenatal diagnosis, infectious diseases, and organ transplantation. Mutation enrichment techniques that allow reduction of unwanted DNA to enable the detection of low-level mutations have emerged since the early 1990s. They are continuously being refined and updated with new technologies. The burgeoning interest in liquid biopsies for residual cancer monitoring, detection of resistance to therapy, and early cancer detection has driven an expanded interest in new and improved methodologies for practical and effective mutation enrichment and detection of low-level mutations of clinical relevance. Content Newly developed mutation enrichment technologies are described and grouped according to the main principle of operation, PCR-blocking technologies, enzymatic methods, and physicochemical approaches. Special emphasis is given to technologies enabling pre-PCR blockage of wtDNA to bypass PCR errors [nuclease-assisted minor-allele enrichment assay with overlapping probes (NaME-PrO) and UV-mediated cross-linking minor allele enrichment (UVME)] or providing high multiplexity followed by next-generation sequencing [Minor allele enriched sequencing through recognition oligonucleotides (MAESTRO)]. Summary This review summarizes technological developments in rare mutation enrichment over the last 12 years, complementing pre-2010 reviews on this topic. The expanding field of liquid biopsy calls for improved limits of detection (LOD) and highly parallel applications, along with the traditional requirements for accuracy, speed, and cost-effectiveness. The current technologies are reviewed with regards to these new requirements.

中文翻译：

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突变富集和检测技术的最新发展

背景 过量的未改变的野生型 DNA (wtDNA) 的存在提供了几乎没有临床价值的信息，通常可能掩盖包含重要诊断或治疗线索的低水平突变。这是生物技术和医学中反复出现的障碍，包括癌症、产前诊断、传染病和器官移植。自 1990 年代初以来，出现了允许减少不需要的 DNA 以检测低水平突变的突变富集技术。它们不断被新技术改进和更新。液体活检用于残余癌症监测、治疗耐药性检测、早期癌症检测已引起人们对新的和改进的方法的兴趣扩大，这些方法用于实用和有效的突变富集和检测临床相关的低水平突变。内容 对新开发的突变富集技术按照主要工作原理、PCR阻断技术、酶法和物理化学方法进行了描述和分组。特别强调了能够对 wtDNA 进行 PCR 前阻断以绕过 PCR 错误的技术 [使用重叠探针 (NaME-PrO) 和 UV 介导的交联次要等位基因富集 (UVME) 的核酸酶辅助次要等位基因富集测定] 或提供高多重性，然后是下一代测序 [通过识别寡核苷酸 (MAESTRO) 进行的次要等位基因富集测序]。总结 本综述总结了过去 12 年中罕见突变富集的技术发展，补充了 2010 年前对该主题的综述。液体活检领域的扩展要求提高检测限 (LOD) 和高度并行的应用，以及对准确性、速度和成本效益的传统要求。针对这些新要求对当前技术进行了审查。