Methylation of microRNAs (miRNAs) is a post-transcriptional modification that affects miRNA activity by altering the specificity of miRNAs to target mRNAs. Abnormal methylation of miRNAs in cancer suggests their potential as a tumor marker. However, the traditional methylated miRNA detection mainly includes mass spectrometry, sequencing and others; complex procedures and reliance on large instruments greatly limit their application in point-of-care testing (POCT). Based on this, we developed DNAzyme-RCA-based gold nanoparticle (AuNP) colorimetric and lateral flow dipstick (LFD) assays to achieve convenient detection of exosomal 5-methylcytosine miRNA-21 (m5C-miRNA-21) for the first time. The two assays achieved specific recognition and linear amplification of m5C-miRNA-21 through the DNAzyme triggered RCA reaction and color output with low background interference through AuNP aggregation induced by base complementary pairing. The lowest concentration of m5C-miRNA-21 visible to the naked eye of the two assays can reach 1 pM and 0.1 pM, respectively. Detection of exosomal m5C-miRNA-21 in clinical blood samples showed that the expression level of m5C-miRNA-21 in colorectal cancer patients was significantly higher than that in healthy individuals. This approach not only demonstrates a new strategy for the detection of colorectal cancer but also provides a reference for the development of novel diagnostic tools for other miRNA methylation-related diseases.

中文翻译：

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基于 DNAzyme-RCA 的比色和侧流试纸测定，用于外泌体 m5C-miRNA-21 的即时检测


microRNA (miRNA) 的甲基化是一种转录后修饰，通过改变 miRNA 对靶标 mRNA 的特异性来影响 miRNA 活性。癌症中 miRNA 的异常甲基化表明它们作为肿瘤标志物的潜力。然而，传统的甲基化miRNA检测主要包括质谱、测序等；复杂的程序和对大型仪器的依赖极大地限制了它们在即时检测（POCT）中的应用。基于此，我们开发了基于DNAzyme-RCA的金纳米颗粒（AuNP）比色法和侧流试纸（LFD）检测方法，首次实现了外泌体5-甲基胞嘧啶miRNA-21（m5C-miRNA-21）的便捷检测。这两种检测通过 DNAzyme 触发 RCA 反应实现了 m5C-miRNA-21 的特异性识别和线性扩增，并通过碱基互补配对诱导的 AuNP 聚集实现了低背景干扰的颜色输出。两次检测中肉眼可见的 m5C-miRNA-21 最低浓度分别可达 1 pM 和 0.1 pM。对临床血液样本中外泌体m5C-miRNA-21的检测表明，结直肠癌患者中m5C-miRNA-21的表达水平显着高于健康个体。该方法不仅展示了检测结直肠癌的新策略，而且为其他miRNA甲基化相关疾病的新型诊断工具的开发提供了参考。