The ability to tailor alterations in genomes, including plant genomes, in a site-specific manner has been greatly advanced through approaches that reduced the complexity and time of genome sequencing along with development of gene editing technologies. These technologies provide a valuable foundation for studies of gene function, metabolic engineering, and trait modification for crop improvement. Development of genome editing methodologies began ∼20 years ago, first with meganucleases and followed by zinc finger nucleases, transcriptional activator-like effector nucleases and, most recently, clustered regulatory interspaced short palindromic repeat (CRISPR)-associated protein (Cas) (CRISPR/Cas), which is by far the most utilized method. The premise of CRISPR/Cas centers on the cleaving of one or both DNA strands by a Cas protein, an endonuclease, followed by mending of the DNA by repair mechanisms inherent in cells. Its user-friendly construct design, greater flexibility in targeting genomic regions, and cost-effective attributes have resulted in it being widely adopted and revolutionizing precise modification of the genomes of many organisms. Indeed, the CRISPR/Cas system has been utilized for gene editing in many plant species, including important food crops, such as maize, wheat, rice, and potatoes. This review summarizes the various approaches, including the most recent designs being used to make modifications from as small as a single-base-pair change to insertion of DNA fragments. On the gene expression level, strategies are presented that make it possible to knock out or modulate through activation and repression. Also discussed are prerequisites necessary for CRISPR/Cas-mediated editing as well as the current challenges.

中文翻译：

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应用基因编辑对植物进行精确的基因修饰。

随着基因编辑技术的发展，通过降低基因组测序的复杂性和时间的方法，以特定位点的方式定制基因组（包括植物基因组）改变的能力已经大大提高。这些技术为基因功能、代谢工程和作物改良性状修饰的研究提供了宝贵的基础。基因组编辑方法的开发始于约 20 年前，首先是大范围核酸酶，随后是锌指核酸酶、类转录激活子效应核酸酶，以及最近的簇状调控间隔短回文重复序列 (CRISPR) 相关蛋白 (Cas) (CRISPR/ Cas)，这是迄今为止使用最多的方法。CRISPR/Cas 的前提是通过 Cas 蛋白（一种核酸内切酶）切割一条或两条 DNA 链，然后通过细胞固有的修复机制修复 DNA。其用户友好的构建设计、靶向基因组区域的更大灵活性以及成本效益的属性使其被广泛采用，并彻底改变了许多生物体基因组的精确修饰。事实上，CRISPR/Cas 系统已被用于许多植物物种的基因编辑，包括重要的粮食作物，如玉米、小麦、水稻和马铃薯。这篇综述总结了各种方法，包括用于进行修改的最新设计，从单碱基对的小改变到 DNA 片段的插入。在基因表达水平上，提出了一些策略，可以通过激活和抑制来敲除或调节。还讨论了 CRISPR/Cas 介导的编辑所需的先决条件以及当前的挑战。