Detection of on-target and off-target mutations generated by CRISPR/Cas9 and other sequence-specific nucleases

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• 作者：Julia Zischewski^a ; Rainer Fischer^a ; ^b ; Luisa Bortesi^a ; ^{luisa.bortesi@molbiotech.rwth-aachen.de}
• 关键词：CRISPR ; Cas9 ; Genome editing ; Engineered nucleases ; Indel detection ; Unbiased off-target analysis ; Mismatch detection assays
• 刊名：Biotechnology Advances
• 出版年：2017
• 出版时间：January-February 2017
• 年：2017
• 卷：35
• 期：1
• 页码：95-104
• 全文大小：508 K
• 卷排序：35

文摘

The development of customizable sequence-specific nucleases such as TALENs, ZFNs and the powerful CRISPR/Cas9 system has revolutionized the field of genome editing. The CRISPR/Cas9 system is particularly versatile and has been applied in numerous species representing all branches of life. Regardless of the target organism, all researchers using sequence-specific nucleases face similar challenges: confirmation of the desired on-target mutation and the detection of off-target events. Here, we evaluate the most widely-used methods for the detection of on-target and off-target mutations in terms of workflow, sensitivity, strengths and weaknesses.