液滴Cas13a-RPA测量揭示了血浆circWDR37在结直肠癌中的潜在作用

Aggregate Pub Date : 2024-09-11 DOI:10.1002/agt2.663
Jingsong Xu, Li Cao, Shuang Yang, Ying Jian, Yu Liu, Zhen Shen, Qian Liu, Xiang Chen, Min Li, Shun Li, Xiaolei Zuo, Min Li, Hua Wang
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引用次数: 0

摘要

大肠癌(CRC)是最常见的癌症之一。循环 RNA 已成为用于癌症诊断和预后评估的有前途的生物标志物。然而,作为 CRC 潜在生物标志物的新型 circRNAs 仍需进一步探索和验证,精确的检测方法也有待开发。在此,我们首次报道了利用液滴Cas13a检测作为CRC生物标志物的circWDR37。通过阵列星 circRNA 微阵列检测、体外和体内功能实验以及 qPCR,我们发现并验证了 circWDR37 是一种用于早期筛查和评估 CRC 预后的生物标记物。通过将微流控液滴与CRISPR/Cas13a和重组酶聚合酶扩增(RPA)相结合,开发出了一种能准确检测circWDR37的新技术--μDCR。同时,还揭示了拥挤剂在提高 Cas13a 性能方面的作用。4% 聚乙二醇 8000 和 3% 右旋糖酐-10 能显著提高一锅 Cas13a-RPA 反应的反应速度和灵敏度。μDCR对circWDR37的检测限为10拷贝/毫升,高于qPCR。临床样本研究结果表明,circWDR37检测可用于有效筛查早期CRC并准确评估预后。CircWDR37 被证实是一种开创性的生物标记物,可用于 CRC 患者的诊断和预后评估。此外,我们用于检测 circWDR37 的创新 µDCR 方法具有操作简便、快速和高通量等显著特点,是无创筛查 CRC 的最佳技术平台。
本文章由计算机程序翻译,如有差异,请以英文原文为准。

Droplets Cas13a‐RPA measurement delineates potential role for plasma circWDR37 in colorectal cancer

Droplets Cas13a‐RPA measurement delineates potential role for plasma circWDR37 in colorectal cancer
Colorectal cancer (CRC) is one of the most prevalent forms of cancer. CircRNAs have emerged as promising biomarkers for cancer diagnosis and prognosis evaluation. However, novel circRNAs as potential biomarkers for CRC still need further exploration and validation, and precise detection methods are yet to be developed. Herein, we report for the first time the use of droplets Cas13a to detect the circWDR37 as a biomarker of CRC. The arraystar circRNA microarray assays, functional experiments in vitro and in vivo, and qPCR were performed to discover and validate that circWDR37 is a biomarker for early screening and prognosis evaluation of CRC. A new technology named µDCR, which accurately detects circWDR37, has been developed by combining microfluidic droplets with CRISPR/Cas13a and recombinase polymerase amplification (RPA). Meanwhile, the role of crowding agent in improving the performance of Cas13a was uncovered. The 4% polyethylene glycol 8000 and 3% dextran‐10 significantly improved the response speed and sensitivity of one‐pot Cas13a‐RPA reaction. The detection limit of circWDR37 by µDCR was found to be 10 copies/mL, which is higher than that of qPCR. The clinical sample findings demonstrated that circWDR37 detection can be utilized to effectively screen for CRC at an early stage and enable accurate assessment of prognosis. CircWDR37 is confirmed as a groundbreaking biomarker for both diagnosis and prognosis evaluation in CRC patients. Furthermore, our innovative µDCR method for detecting circWDR37 demonstrates impressive attributes such as streamlined operation, rapidity, and high‐throughput, making it an optimal technology platform for the noninvasive screening of CRC.
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