A smart polymer for sequence-selective binding, pulldown, and release of DNA targets. A smart polymer for sequence-selective binding, pulldown, and release of DNA targets

Selective isolation of DNA is crucial for applications in biology, bionanotechnology, clinical diagnostics and forensics. We herein report a smart methanol-responsive polymer (MeRPy) that can be programmed to bind and separate single- as well as double-stranded DNA targets. Captured targets are quickly isolated and released back into solution by denaturation (sequence-agnostic) or toehold-mediated strand displacement (sequence-selective). The latter mode allows 99.8% efficient removal of unwanted sequences and 79% recovery of highly pure target sequences. We applied MeRPy for the depletion of insulin, glucagon, and transthyretin cDNA from clinical next-generation sequencing (NGS) libraries. This step improved data quality for low-abundance transcripts in expression profiles of pancreatic tissues. Its low cost, scalability, high stability and ease of use make MeRPy suitable for diverse applications in research and clinical laboratories, including enhancement of NGS libraries, extraction of DNA from biological samples, preparative-scale DNA isolations, and sorting of DNA-labeled non-nucleic acid targets. Overall design: 9 Samples are analyzed: 3 untreated samples that were not depleted with MeRPy; 3 negative control samples that were treated with MeRPy but lacked the target-specific catcher strand library (CSL); 3 experiment samples that were treated with MeRPy and a target-specific CSL.

DNA的选择性分离在生物学、生物纳米技术、临床诊断及法医学领域均具有至关重要的应用价值。
本文报道了一种智能甲醇响应聚合物（methanol-responsive polymer，MeRPy），该材料可经编程实现对单链及双链DNA靶标的结合与分离。捕获得到的靶标可通过变性（不依赖序列）或toehold介导的链置换（toehold-mediated strand displacement）方式快速解离并释放回溶液中。其中，后一种解离模式可实现99.8%的非靶序列去除率与79%的高纯度靶序列回收率。
我们将MeRPy应用于临床二代测序（next-generation sequencing，NGS）文库中胰岛素、胰高血糖素及甲状腺素转运蛋白cDNA的消减。该步骤显著提升了胰腺组织表达谱中低丰度转录本的测序数据质量。
MeRPy具备成本低廉、可规模化制备、稳定性优异且操作简便等优势，适用于科研与临床实验室的多种应用场景，包括NGS文库质量优化、生物样本DNA提取、制备级DNA分离以及DNA标记非核酸靶标的分选等。
实验整体设计：共分析9份样本：3份未经MeRPy处理的空白对照样本；3份经MeRPy处理但未添加靶标特异性捕获链文库（catcher strand library，CSL）的阴性对照样本；以及3份经MeRPy与靶标特异性CSL共同处理的实验组样本。