Methods for Optimizing CRISPR-Cas9 Genome Editing Specificity.
Methods for Optimizing CRISPR-Cas9 Genome Editing Specificity.
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DOI：
10.1016/j.molcel.2016.07.004
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发表时间：
2016-08-04
期刊：
影响因子：
16
通讯作者：
Hsu PD
中科院分区：
文献类型：
--
作者：
Tycko J;Myer VE;Hsu PD
CRISPR-Cas9基因组工程工具箱在传递、修复和特异性策略方面的进展正在帮助研究人员以前所未有的精度和灵敏度了解基因功能。CRISPR-Cas9还具有通过直接纠正致病突变来治疗遗传疾病的巨大治疗潜力。虽然Cas9蛋白已被证明在非靶点结合和切割DNA，但随着引导RNA选择、蛋白质和引导工程、新型酶和非靶点检测方法的显著改进，Cas9特异性领域正在迅速发展。我们回顾了该领域的重要挑战和突破，作为对基因组编辑技术感兴趣的用户的全面实践指南，强调了优化特异性的关键工具和战略。基因组编辑界现在应该努力标准化这种测量和报告非靶标活动的方法，同时记住，特异性的目标应该是继续改进和警惕。
Advances in the development of delivery, repair, and specificity strategies for the CRISPR-Cas9 genome engineering toolbox are helping researchers understand gene function with unprecedented precision and sensitivity. CRISPR-Cas9 also holds enormous therapeutic potential for the treatment of genetic disorders by directly correcting disease-causing mutations. Although the Cas9 protein has been shown to bind and cleave DNA at off-target sites, the field of Cas9 specificity is rapidly progressing with marked improvements in guide RNA selection, protein and guide engineering, novel enzymes, and off-target detection methods. We review important challenges and breakthroughs in the field as a comprehensive practical guide to interested users of genome editing technologies, highlighting key tools and strategies for optimizing specificity. The genome editing community should now strive to standardize such methods for measuring and reporting off-target activity, while keeping in mind that the goal for specificity should be continued improvement and vigilance.