Zhang Kun, Li Yan, Jiang Shengjie, Ju Shang
Wound Treatment Center, Xinxiang Central Hospital, The Fourth Clinical College of Xinxiang Medical University, Xinxiang, Henan 453000, China.
Xinxiang Key Laboratory on Healing Mechanism Research of Diabetic Foot Ulcer, Xinxiang Central Hospital, The Fourth Clinical College of Xinxiang Medical University, Xinxiang, Henan 453000, China.
ACS Omega. 2024 Feb 23;9(9):10897-10903. doi: 10.1021/acsomega.3c10003. eCollection 2024 Mar 5.
MicroRNAs (miRNAs) have significant regulatory functions in the modulation of gene expression, making them essential biomarkers for the diagnosis and prognosis of diseases. Nevertheless, the identification of miRNA poses significant difficulty in terms of its low abundance, necessitating sensitive and reliable approaches. Herein, we develop a simple approach, termed atalytic assembly of NAzyme ntegrates with rimer xchange eaction (CDiPER), for reliable and sensitive miRNA detection through the target recognition-triggered DNAzyme assembly and primer exchange reaction (PER) strategy. In this method, target miRNA can precisely bind with a specifically designed hairpin probe (H probe) to induce the conformation changes of the H probe, releasing DNAzyme sections to activate the PER process for signal amplification and fluorescence signal production. The established method displays a high dynamic range of over 6 orders of magnitude and a low detection limit of 312 aM. The created method has a number of unique advantages, such as (i) a better sensitivity than existing systems using PER for signal amplification as a result of its integration with the target recognition-triggered DNAzyme assembly and (ii) streamlined operating procedures. Further, the technology was used to detect the expression of miRNA in collected clinical samples from diabetes mellitus patients, revealing that miRNA was decreased in patients and demonstrating the significant clinical promise of the method.
微小RNA（miRNA）在基因表达调控中具有重要的调节功能，使其成为疾病诊断和预后的重要生物标志物。然而，由于miRNA丰度低，其鉴定存在重大困难，因此需要灵敏且可靠的方法。在此，我们开发了一种简单的方法，称为核酸酶催化组装与引物交换反应相结合（CDiPER），通过靶标识别触发的DNA酶组装和引物交换反应（PER）策略，实现对miRNA的可靠且灵敏的检测。在该方法中，靶标miRNA可与特异性设计的发夹探针（H探针）精确结合，诱导H探针的构象变化，释放DNA酶片段以激活PER过程进行信号放大和产生荧光信号。所建立的方法显示出超过6个数量级的高动态范围和312 aM的低检测限。该方法具有许多独特优势，例如：（i）由于其与靶标识别触发的DNA酶组装相结合，比现有的使用PER进行信号放大的系统具有更高的灵敏度；（ii）操作流程简化。此外，该技术用于检测糖尿病患者收集的临床样本中miRNA的表达，结果显示患者体内miRNA水平降低，证明了该方法具有重大的临床应用前景。
