Abstract：
The clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 system has been widely used to create animal models for biomedical and agricultural use owing to its low cost and easy handling. However, the occurrence of erroneous cleavage (off-targeting) may raise certain concerns for the practical application of the CRISPR-Cas9 system. In this study, we created a melanocortin 1 receptor (MC1R)-edited pig model through somatic cell nuclear transfer (SCNT) by using porcine kidney cells modified by the CRISPR-Cas9 system. We then carried out whole-genome sequencing of two MC1R-edited pigs and two cloned wild-type siblings, together with the donor cells, to assess the genome-wide presence of single-nucleotide variants and small insertions and deletions (indels) and found only one candidate off-target indel in both MC1R-edited pigs. In summary, our study indicates that the minimal off-targeting effect induced by CRISPR-Cas9 may not be a major concern in gene-edited pigs created by SCNT.
摘要：
成簇的规则间隔短回文重复序列（CRISPR）-Cas9系统由于其低成本和易于处理而被广泛用于创建用于生物医学和农业用途的动物模型。然而,错误切割(脱靶)的发生可能会引起CRISPR-Cas9系统实际应用的某些担忧.在这项研究中,我们使用CRISPR-Cas9系统修饰的猪肾细胞，通过体细胞核移植（SCNT）创建了黑皮质素1受体（MC1R）编辑的猪模型。然后，我们对两只MC1R编辑的猪和两只克隆的野生型兄弟姐妹进行了全基因组测序，连同供体细胞，评估全基因组存在的单核苷酸变异和小的插入和缺失（indel），在两只MC1R编辑的猪中只有一个候选脱靶indel。总之,我们的研究表明,CRISPR-Cas9诱导的最小脱靶效应可能不是SCNT产生的基因编辑猪的主要关注点.
