Abstract

A photoactivatable CRISPR/Cas9 system consisting of the Cas9 protein, synthetic 102-nt sgRNA or a pair of guide crRNA/tracrRNA, and blocking photocleavable oligodeoxyribonucleotides has been designed. Photocleavable oligodeoxyribonucleotides contain two or three photodegradable linkers in their structure. The thermal stability of complexes of these modified oligonucleotides with guide crRNA decreases with increasing the number of linkers. The kinetics of the photomodified oligodeoxyribonucleotides cleavage has been investigated. It has been shown that UV irradiation of the CRISPR/Cas9 system in the presence of photomodified blocking oligodeoxyribonucleotides can result in photoactivatable gene editing in vitro.

中文翻译：

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可光激活的CRISPR / Cas9系统

摘要

设计了一种光活化的CRISPR / Cas9系统，该系统由Cas9蛋白，合成的102-nt sgRNA或一对引导性crRNA / tracrRNA组成，并能阻断可光裂解的寡脱氧核糖核苷酸。可光裂解的寡脱氧核糖核苷酸在其结构中包含两个或三个可光降解的接头。这些修饰寡核苷酸与向导crRNA的复合物的热稳定性随连接子数目的增加而降低。已经研究了光改性的寡脱氧核糖核苷酸裂解的动力学。已经显示，在存在光修饰的封闭寡聚脱氧核糖核苷酸的情况下，CRISPR / Cas9系统的紫外线照射可导致体外可光激活的基因编辑。