The polymerase chain reaction (PCR) plays a central role in genetic engineering and is routinely used in various applications, from biological and medical research to the diagnosis of viral infections. PCR is an extremely sensitive method for detecting target DNA sequences, but it is substantially error prone. In particular, the mishybridization of primers to contaminating sequences can result in false positives for virus tests. The blocker method, also called the clamping method, has been developed to suppress mishybridization errors. However, its application is limited by the requirement that the contaminating template sequence be known in advance. Here, we demonstrate that a mixture of multiple blocker sequences effectively suppresses the amplification of contaminating sequences even in the presence of uncertainty. The blocking effect was characterized by a simple model validated by experiments. Furthermore, the modeling allowed us to minimize the errors by optimizing the blocker concentrations. The results highlighted an inherent robustness of the blocker method in that fine-tuning the blocker concentrations is not necessary. Our method extends the applicability of PCR and other hybridization-based techniques, including genome editing, RNA interference, and DNA nanotechnology, by improving their fidelity.

中文翻译：

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阻断 PCR 的不确定错误


聚合酶链反应 （PCR） 在基因工程中起着核心作用，通常用于各种应用，从生物学和医学研究到病毒感染的诊断。PCR 是一种检测靶标 DNA 序列的极其灵敏的方法，但它极易出错。特别是，引物与污染序列的错误杂交会导致病毒检测的假阳性。阻塞方法，也称为夹紧方法，已被开发用于抑制错误杂交错误。但是，其应用受到事先知道污染模板序列的要求的限制。在这里，我们证明即使在存在不确定性的情况下，多个阻断序列的混合物也能有效抑制污染序列的扩增。阻断效应通过一个经过实验验证的简单模型来表征。此外，该建模使我们能够通过优化阻滞剂浓度来最大限度地减少误差。结果突出了封闭剂方法固有的稳定性，因为无需微调封闭剂浓度。我们的方法通过提高 PCR 和其他基于杂交的技术（包括基因组编辑、RNA 干扰和 DNA 纳米技术）的保真度，扩展了它们的适用性。