Single-nucleotide editing for zebra3 and wsl5 phenotypes in rice using CRISPR/Cas9-mediated adenine base editors

作     者：Kutubuddin A.Molla Justin Shih Yinong Yang Kutubuddin A.Molla;Justin Shih;Yinong Yang

作者机构：Department of Plant Pathology and Environmental Microbiology and the Huck Institute of the Life SciencesThe Pennsylvania State UniversityUniversity ParkPA 16802USA ICAR-National Rice Research InstituteCuttack 753006India 

出 版 物：《aBIOTECH》 (生物技术通报（英文版）)

年 卷 期：2020年第1卷第2期

页      面：106-118页

核心收录：

学科分类：0710[理学-生物学] 07[理学] 09[农学] 071007[理学-遗传学] 0901[农学-作物学] 090102[农学-作物遗传育种] 

基　　金：This work was supported by National Science Foundation Plant Genome Research Program Grant No.1740874 the USDA National Institute of Food and Agriculture and Hatch Appropriations under Project#PEN04659 and Accession#1016432 to Yinong Yang 

主　　题：Adenine base editor CRISPR/Cas9 genome editing Plant base editing Precise point mutation Transgene-free rice 

摘      要：The CRISPR/Cas9-mediated base editing technology can efficiently generate point mutations in the genome without introducing a double-strand break(DSB)or supplying a DNA donor template for homology-directed repair(HDR).In this study,adenine base editors(ABEs)were used for rapid generation of precise point mutations in two distinct genes,OsWSL5,and OsZEBRA3(Z3),in both rice protoplasts and regenerated *** precisely engineered point mutations were stably inherited to subsequent *** single nucleotide alterations resulted in single amino acid changes and associated wsl5 and z3 phenotypes as evidenced by white stripe leaf and light green/dark green leaf pattern,*** selfing and genetic segregation,transgene-free,base edited wsl5 and z3 mutants were obtained in a short period of *** noticed a novel mutation(V540A)in Z3 locus could also mimic the phenotype of Z3 mutation(S542P).Furthermore,we observed unexpected non-A/G or T/C mutations in the ABE editing window in a few of the edited *** ABE vectors and the method from this study could be used to simultaneously generate point mutations in multiple target genes in a single transformation and serve as a useful base editing tool for crop improvement as well as basic studies in plant biology.