Development of Single Molecule Techniques for Sensing and Manipulation of CRISPR and Polymerase Enzymes.
Development of Single Molecule Techniques for Sensing and Manipulation of CRISPR and Polymerase Enzymes.
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DOI：
10.1002/smll.202300328
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发表时间：
2023-05
期刊：
影响因子：
13.3
通讯作者：
Jose Chu;Andres Romero;Jeffrey Taulbee;Kiana Aran
中科院分区：
文献类型：
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作者：
Jose Chu;Andres Romero;Jeffrey Taulbee;Kiana Aran
规则间隔短回文重复序列（CRISPR）和聚合酶是强大的酶，它们在基因组学、蛋白质组学和转录组学中的多种应用彻底改变了当今的生物技术产业。CRISPR已被广泛用于基因组编辑应用，聚合酶可以通过聚合酶链反应（PCR）有效地扩增基因组转录物。对这些酶的进一步研究可以揭示其机制的具体细节，从而大大扩展其用途。单分子技术是探测酶机制的有效方法，因为它们可以比整体或批量生物传感技术更详细地解析中间构象和状态。这篇综述讨论了传感和操纵单个生物分子的各种技术，可以帮助促进和加快这些发现。每个平台都分为光学、机械或电子平台。简要介绍了每种技术的方法，操作原理，输出和效用，然后讨论了它们在单分子水平上监测和控制CRISPR和聚合酶的应用，并简要概述了它们的局限性和未来前景。
Clustered regularly interspaced short palindromic repeats (CRISPR) and polymerases are powerful enzymes and their diverse applications in genomics, proteomics, and transcriptomics have revolutionized the biotechnology industry today. CRISPR has been widely adopted for genomic editing applications and Polymerases can efficiently amplify genomic transcripts via polymerase chain reaction (PCR). Further investigations into these enzymes can reveal specific details about their mechanisms that greatly expand their use. Single-molecule techniques are an effective way to probe enzymatic mechanisms because they may resolve intermediary conformations and states with greater detail than ensemble or bulk biosensing techniques. This review discusses various techniques for sensing and manipulation of single biomolecules that can help facilitate and expedite these discoveries. Each platform is categorized as optical, mechanical, or electronic. The methods, operating principles, outputs, and utility of each technique are briefly introduced, followed by a discussion of their applications to monitor and control CRISPR and Polymerases at the single molecule level, and closing with a brief overview of their limitations and future prospects.