The discovery and function analysis of terminal deoxynucleotidyl transferase (TdT) add a new dimension to the understanding of leukemia mechanisms and stimulate the development of new analytical tools for leukemia diagnosis. Herein, taking advantage of the inherent property of TdT for performing DNA synthesis using only single-stranded DNA as the nucleic acid substrate, we developed a self-customized multichannel exponential amplification (SMEA) system for the fluorescent sensing of TdT activity. The SMEA design employs an intermolecular DNA interaction made of a nicking site-incorporated elongation primer (EP) and a nicking site-incorporated poly-thymine tailed molecular beacon (Poly-T-MB). The absence of TdT is unable to bridge the relationship between EP and Poly-T-MB, ensuring the SMEA has an ultralow background. The presence of TdT, however, leads to the elongation of EP to poly-adenine tailed EP (Poly-A-EP) under a dATP pool responsible for further hybridization with numerous Poly-T-MB. With the aid of polymerase and nickase to react with the hybridization product of Poly-A-EP/(Poly-T-MB)_n, it can cause bidirectional strand nicking, polymerization, and displacement in many cycles and channels. In this case, the SMEA is found to be associated with the configuration transformation and splitting of all Poly-T-MBs for a significant fluorescence enhancement. Depending on this high target signal amplification and strong background inhibition abilities, the SMEA sensing system is powerful for the qualitative and quantitative determination of TdT activity, showing that it has great promise for biomedical study and disease diagnosis.

中文翻译：

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自行定制的多通道指数扩增调节末端脱氧核苷酸转移酶活性的动力监测

末端脱氧核苷酸转移酶（TdT）的发现和功能分析为白血病机制的理解增加了一个新的维度，并刺激了白血病诊断新分析工具的发展。在此，利用 TdT 的固有特性，仅使用单链 DNA 作为核酸底物进行 DNA 合成，我们开发了一种用于 TdT 活性荧光传感的自定义多通道指数放大 (SMEA) 系统。SMEA 设计采用分子间 DNA 相互作用，由结合切口位点的延伸引物 (EP) 和结合切口位点的多胸腺嘧啶尾分子信标 (Poly-T-MB) 组成。TdT 的缺失无法弥合 EP 和 Poly-T-MB 之间的关系，从而确保 SMEA 具有超低背景。然而，TdT 的存在，导致 EP 在 dATP 池下延伸为聚腺嘌呤尾 EP (Poly-A-EP)，负责与许多 Poly-T-MB 进一步杂交。借助聚合酶和切口酶与Poly-A-EP/(Poly-T-MB)的杂交产物反应_n，它可以在许多循环和通道中引起双向链切口、聚合和位移。在这种情况下，发现 SMEA 与所有 Poly-T-MB 的配置转换和分裂相关，从而显着增强荧光。凭借这种高目标信号放大和强大的背景抑制能力，SMEA传感系统对TdT活性的定性和定量测定具有强大的功能，表明它在生物医学研究和疾病诊断方面具有很大的应用前景。