Cancer remains a formidable global health challenge, necessitating precise diagnostic tools. The KRAS gene, particularly the G12D mutation, holds pivotal clinical significance across various cancer subtypes. Existing DNA sensing techniques using single-stranded and double-stranded oligonucleotide probes are susceptible for enzymatic degradation, especially in complex biological samples like blood or serum. To address these challenges, we present an innovative approach that involves tuning assembled DNA nanoarchitecture into a disassembled state to accurate and ultrasensitive detection of the KRAS G12D mutation. Our method utilized a pair of multifunctional assembly hairpin probes to construct a DNA wire with both of a fluorophore labeled probe and a quencher labeled probe attached. Upon binding with the target DNA, the DNA wire undergoes gradual disassembly, separating all fluorophores from quenchers to significantly amplify fluorescence signals. This approach offers exceptional sensitivity and reaction kinetics, capable of detecting the KRAS G12D at femtomolar levels, and demonstrates outstanding specificity in discriminating single-base mutations. Furthermore, the DNA nanoarchitecture with well-demonstrated ability to nuclease resistance makes our method exhibits robustness in detecting KRAS G12D mutation in human serum samples. This innovative approach represents an advancement in cancer diagnostics, providing a promising tool for early disease diagnosis and biomedical study.

中文翻译：

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将组装的 DNA 纳米结构调整为分解状态，以增强 KRAS G12D 突变的准确和超灵敏检测


癌症仍然是一个巨大的全球健康挑战，需要精确的诊断工具。 KRAS 基因，特别是 G12D 突变，在各种癌症亚型中具有关键的临床意义。现有的使用单链和双链寡核苷酸探针的 DNA 传感技术很容易被酶促降解，尤其是在复杂的生物样品（如血液或血清）中。为了应对这些挑战，我们提出了一种创新方法，包括将组装的 DNA 纳米结构调整为分解状态，以准确、超灵敏地检测 KRAS G12D 突变。我们的方法利用一对多功能组装发夹探针来构建连接有荧光团标记探针和猝灭剂标记探针的DNA线。与目标 DNA 结合后，DNA 线逐渐分解，将所有荧光团与猝灭剂分离，从而显着放大荧光信号。这种方法提供了卓越的灵敏度和反应动力学，能够检测飞摩尔水平的 KRAS G12D，并在区分单碱基突变方面表现出出色的特异性。此外，DNA纳米结构具有良好的核酸酶抗性，使我们的方法在检测人血清样本中的KRAS G12D突变方面表现出稳健性。这种创新方法代表了癌症诊断的进步，为早期疾病诊断和生物医学研究提供了有前景的工具。