Telomerase is considered a biomarker for the early diagnosis and clinical treatment of cancer. The rapid and sensitive detection of telomerase activity is crucial to biological research, clinical diagnosis, and drug development. However, the main obstacles facing the current telomerase activity assay are the cumbersome and time-consuming procedure, the easy degradation of the telomerase RNA template and the need for additional proteases. Therefore, it is necessary to construct a new method for the detection of telomerase activity with easy steps, efficient reaction and strong anti-interference ability. Herein, an efficient, enzyme-free, economical, sensitive, fluorometric detection method for telomerase activity in one-step, named triggered-DNA (T-DNA) nanomachine, was created based on target-triggered DNAzyme-cleavage activity and catalytic molecular beacon (CMB). Telomerase served as a switch and extended few numbers of (TTAGGG) repeat sequences to initiate the signal amplification in the T-DNA nanomachine, resulting in a strong fluorescent signal. The reaction was a one-step method with a shortened time of 1 h and a constant temperature of 37 °C, without the addition of any protease. It also sensitively distinguished telomerase activity in various cell lines. The T-DNA nanomachine offered a detection limit of 12 HeLa cells μL, 9 SK-Hep-1 cells μL and 3 HuH-7 cells μL with a linear correlation detection range of 0.39 × 10–6.25 × 10 HeLa cells μL for telomerase activity. In conclusion, our study demonstrated that the triggered-DNA nanomachine fulfills the requirements for rapid detection of telomerase activity in one-step under isothermal and enzyme-free conditions with excellent specificity, and its simple and stable structure makes it ideal for complex systems. These findings indicated the application prospect of DNA nanomachines in clinical diagnostics and provided new insights into the field of DNA nanomachine-based bioanalysis.

中文翻译：

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一种无酶触发DNA纳米机器，用于一步法快速检测端粒酶活性

端粒酶被认为是癌症早期诊断和临床治疗的生物标志物。快速、灵敏地检测端粒酶活性对于生物学研究、临床诊断和药物开发至关重要。然而，目前端粒酶活性测定面临的主要障碍是程序繁琐且耗时、端粒酶RNA模板容易降解以及需要额外的蛋白酶。因此，有必要构建一种步骤简单、反应高效、抗干扰能力强的端粒酶活性检测新方法。在此，基于目标触发DNA酶裂解活性和催化分子信标，创建了一种高效、无酶、经济、灵敏的荧光一步检测端粒酶活性的方法，称为触发DNA（T-DNA）纳米机器。 （中巴）。端粒酶充当开关并延伸少量 (TTAGGG) 重复序列以启动 T-DNA 纳米机器中的信号放大，从而产生强烈的荧光信号。反应为一步法，时间缩短为1 h，恒温37℃，不添加任何蛋白酶。它还灵敏地区分各种细胞系中的端粒酶活性。 T-DNA 纳米机器的端粒酶活性检测限为 12 HeLa 细胞 μL、9 SK-Hep-1 细胞 μL 和 3 HuH-7 细胞 μL，线性相关检测范围为 0.39 × 10–6.25 × 10 HeLa 细胞 μL 。总之，我们的研究表明，触发DNA纳米机器满足在等温无酶条件下一步快速检测端粒酶活性的要求，具有良好的特异性，其简单稳定的结构使其成为复杂系统的理想选择。这些发现揭示了DNA纳米机器在临床诊断中的应用前景，并为基于DNA纳米机器的生物分析领域提供了新的见解。