{"title":"利用液滴数字 CRISPR/Cas13a 平台在护理点定量检测环状 RNA 和 microRNA。","authors":"","doi":"10.1016/j.bios.2024.116825","DOIUrl":null,"url":null,"abstract":"<div><div>Circular RNA (circRNA) and microRNA (miRNA) are both non-coding RNAs (ncRNAs) that serve as biomarkers for cancer diagnosis and prognosis. Quantitative detection of these ncRNAs is of particular importance to elucidate the functional mechanisms and evaluate their potential as biomarkers. However, the inherent structures of circRNA and miRNA are different from the mRNA, conventional qRT-PCR is unsuitable for the detection of these ncRNAs. Here, we propose a sensitive method for quantitative detection of circRNA and miRNA using polydisperse droplet digital CRISPR/Cas13a (PddCas13a). It can achieve limits of detection (LOD) as low as ∼10 aM without polymerase-based amplification. To efficiently detect the circRNA and miRNA in real samples, we use a chemically modified crRNA to enhance the stability of crRNA and improve the performance of Cas13a in complex environments containing contaminants. By integrating an extraction-free procedure with PddCas13a, we experimentally demonstrate the applicability of PddCas13a by testing clinical samples. Furthermore, we develop an automated and portable instrument for PddCas13a and verify its applicability for the detection of circRNA and miRNA from exosomes in point-of-care (POC) setting. This is the first report to detect the circRNA and miRNA simultaneously in POC setting. We envision this platform could promote the research of ncRNAs.</div></div>","PeriodicalId":259,"journal":{"name":"Biosensors and Bioelectronics","volume":null,"pages":null},"PeriodicalIF":10.7000,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Quantitative detection of circular RNA and microRNA at point-of-care using droplet digital CRISPR/Cas13a platform\",\"authors\":\"\",\"doi\":\"10.1016/j.bios.2024.116825\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><div>Circular RNA (circRNA) and microRNA (miRNA) are both non-coding RNAs (ncRNAs) that serve as biomarkers for cancer diagnosis and prognosis. Quantitative detection of these ncRNAs is of particular importance to elucidate the functional mechanisms and evaluate their potential as biomarkers. However, the inherent structures of circRNA and miRNA are different from the mRNA, conventional qRT-PCR is unsuitable for the detection of these ncRNAs. Here, we propose a sensitive method for quantitative detection of circRNA and miRNA using polydisperse droplet digital CRISPR/Cas13a (PddCas13a). It can achieve limits of detection (LOD) as low as ∼10 aM without polymerase-based amplification. To efficiently detect the circRNA and miRNA in real samples, we use a chemically modified crRNA to enhance the stability of crRNA and improve the performance of Cas13a in complex environments containing contaminants. By integrating an extraction-free procedure with PddCas13a, we experimentally demonstrate the applicability of PddCas13a by testing clinical samples. Furthermore, we develop an automated and portable instrument for PddCas13a and verify its applicability for the detection of circRNA and miRNA from exosomes in point-of-care (POC) setting. This is the first report to detect the circRNA and miRNA simultaneously in POC setting. We envision this platform could promote the research of ncRNAs.</div></div>\",\"PeriodicalId\":259,\"journal\":{\"name\":\"Biosensors and Bioelectronics\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":10.7000,\"publicationDate\":\"2024-10-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Biosensors and Bioelectronics\",\"FirstCategoryId\":\"1\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S0956566324008327\",\"RegionNum\":1,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q1\",\"JCRName\":\"BIOPHYSICS\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Biosensors and Bioelectronics","FirstCategoryId":"1","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0956566324008327","RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"BIOPHYSICS","Score":null,"Total":0}
Quantitative detection of circular RNA and microRNA at point-of-care using droplet digital CRISPR/Cas13a platform
Circular RNA (circRNA) and microRNA (miRNA) are both non-coding RNAs (ncRNAs) that serve as biomarkers for cancer diagnosis and prognosis. Quantitative detection of these ncRNAs is of particular importance to elucidate the functional mechanisms and evaluate their potential as biomarkers. However, the inherent structures of circRNA and miRNA are different from the mRNA, conventional qRT-PCR is unsuitable for the detection of these ncRNAs. Here, we propose a sensitive method for quantitative detection of circRNA and miRNA using polydisperse droplet digital CRISPR/Cas13a (PddCas13a). It can achieve limits of detection (LOD) as low as ∼10 aM without polymerase-based amplification. To efficiently detect the circRNA and miRNA in real samples, we use a chemically modified crRNA to enhance the stability of crRNA and improve the performance of Cas13a in complex environments containing contaminants. By integrating an extraction-free procedure with PddCas13a, we experimentally demonstrate the applicability of PddCas13a by testing clinical samples. Furthermore, we develop an automated and portable instrument for PddCas13a and verify its applicability for the detection of circRNA and miRNA from exosomes in point-of-care (POC) setting. This is the first report to detect the circRNA and miRNA simultaneously in POC setting. We envision this platform could promote the research of ncRNAs.
期刊介绍:
Biosensors & Bioelectronics, along with its open access companion journal Biosensors & Bioelectronics: X, is the leading international publication in the field of biosensors and bioelectronics. It covers research, design, development, and application of biosensors, which are analytical devices incorporating biological materials with physicochemical transducers. These devices, including sensors, DNA chips, electronic noses, and lab-on-a-chip, produce digital signals proportional to specific analytes. Examples include immunosensors and enzyme-based biosensors, applied in various fields such as medicine, environmental monitoring, and food industry. The journal also focuses on molecular and supramolecular structures for enhancing device performance.