Whole-genome sequencing reveals rare off-target mutations in CRISPR/Cas9-edited grapevine

作     者：Xianhang Wang Mingxing Tu Ya Wang Wuchen Yin Yu Zhang Hongsong Wu Yincong Gu Zhi Li Zhumei Xi Xiping Wang Xianhang Wang;Mingxing Tu;Ya Wang;Wuchen Yin;Yu Zhang;Hongsong Wu;Yincong Gu;Zhi Li;Zhumei Xi;Xiping Wang

作者机构：State Key Laboratory of Crop Stress Biology in Arid AreasCollege of HorticultureNorthwest A&F University712100 YanglingShaanxiChina College of EnologyNorthwest A&F University712100 YanglingShaanxiChina Key Laboratory of Horticultural Plant Biology and Germplasm Innovation in Northwest ChinaMinistry of AgricultureNorthwest A&F University712100 YanglingShaanxiChina Novogene Technologies Corporation100000 BeijingChina OEbiotech Corporation200000 ShanghaiChina 

出 版 物：《Horticulture Research》 (园艺研究（英文）)

年 卷 期：2021年第8卷第1期

页      面：1352-1362页

核心收录：

学科分类：09[农学] 0902[农学-园艺学] 090201[农学-果树学] 

基　　金：the National Natural Science Foundation of China(U1603234,31572110,and 32002000) the Program for Innovative Research Team of Grape Germplasm Resources and Breeding(2013KCT-25) 

主　　题：Whole Sanger rare 

摘      要：The CRISPR(clustered regularly interspaced short palindromic repeats)-associated protein 9(Cas9)system is a powerful tool for targeted genome editing,with applications that include plant biotechnology and functional genomics ***,the specificity of Cas9 targeting is poorly investigated in many plant species,including fruit *** assess the off-target mutation rate in grapevine(Vitis vinifera),we performed whole-genome sequencing(WGS)of seven Cas9-edited grapevine plants in which one of two genes was targeted by CRISPR/Cas9 and three wild-type(WT)*** total,we identified between 202,008 and 272,397 single nucleotide polymorphisms(SNPs)and between 26,391 and 55,414 insertions/deletions(indels)in the seven Cas9-edited grapevine plants compared with the three WT ***,3272 potential off-target sites were selected for further *** one off-target indel mutation was identified from the WGS data and validated by Sanger *** addition,we found 243 newly generated off-target sites caused by genetic variants between the Thompson Seedless cultivar and the grape reference genome(PN40024)but no true off-target *** conclusion,we observed high specificity of CRISPR/Cas9 for genome editing of grapevine.