Single-nucleotide polymorphisms contribute to phenotypic diversity in maize. Creation and functional annotation of point mutations has been limited by the low efficiency of conventional methods based on random mutatio...Single-nucleotide polymorphisms contribute to phenotypic diversity in maize. Creation and functional annotation of point mutations has been limited by the low efficiency of conventional methods based on random mutation. An efficient tool for generating targeted single-base mutations is desirable for both functional genomics and precise genetic improvement. The objective of this study was to test the efficiency of targeted C-to-T base editing of two non-allelic acetolactate synthase(ALS) in generating sulfonylurea herbicide-resistant mutants. A CRISPR/Cas9 nickase-cytidine deaminase fused with uracil DNA glycosylase inhibitor(UGI) was employed to achieve targeted conversion of cytosine to thymine in ZmALS1 and ZmALS2. Both protoplasts and recovered mutant plants showed the activity of the cytosine base editor, with an in vivo efficiency of up to 13.8%. Transgene-free edited plants harboring a homozygous ZmALS1 mutation or a ZmALS1 and ZmALS2 double mutation were tested for their resistance at a dose of up to 15-fold the recommended limit of chlorsulfuron, a sulfonylurea herbicide widely used in agriculture. Targeted base editing of C-to-T per se and a phenotype verified in the generated mutants demonstrates the power of base editing in precise maize breeding.展开更多
To confirm allelic relationship between Lr9 and the leaf rust resistance gene in KLM4-3B, genetics of resistance was studied using crosses (WL711 + Lr9) × WL711 and (WL711 + LrKLM4-3B) × WL711. The F2 popula...To confirm allelic relationship between Lr9 and the leaf rust resistance gene in KLM4-3B, genetics of resistance was studied using crosses (WL711 + Lr9) × WL711 and (WL711 + LrKLM4-3B) × WL711. The F2 populations in cross (WL711 + Lr9) × WL711 and (WL711 + LrKLM4-3B) × WL711 segregated in ratio of 3:1 for disease reaction at seedling stage against pathotype 77-5 of leaf rust. This suggests that rust resistance in these stocks are under the control of single dominant genes. Further, to study allelic relationship betweenLr 9 and LrKLM4-3B, F2 population of the cross (WL711 + LrKLM4-3B) × (WL711 + Lr9) was studied. A segregation ratio of 15:1 implies that the two genes Lr9 and LrKLM4-3B are non-allelic genes.展开更多
基金supported by the Key Area Research and Development Program of Guangdong Province(2018B020202008)the National Natural Science Foundation of China(31771808)+2 种基金Beijing Municipal Science and Technology Project(D171100007717001)the National Key Research and Development Program of China(2016YFD0101803)National Engineering Laboratory for Crop Molecular Breeding。
文摘Single-nucleotide polymorphisms contribute to phenotypic diversity in maize. Creation and functional annotation of point mutations has been limited by the low efficiency of conventional methods based on random mutation. An efficient tool for generating targeted single-base mutations is desirable for both functional genomics and precise genetic improvement. The objective of this study was to test the efficiency of targeted C-to-T base editing of two non-allelic acetolactate synthase(ALS) in generating sulfonylurea herbicide-resistant mutants. A CRISPR/Cas9 nickase-cytidine deaminase fused with uracil DNA glycosylase inhibitor(UGI) was employed to achieve targeted conversion of cytosine to thymine in ZmALS1 and ZmALS2. Both protoplasts and recovered mutant plants showed the activity of the cytosine base editor, with an in vivo efficiency of up to 13.8%. Transgene-free edited plants harboring a homozygous ZmALS1 mutation or a ZmALS1 and ZmALS2 double mutation were tested for their resistance at a dose of up to 15-fold the recommended limit of chlorsulfuron, a sulfonylurea herbicide widely used in agriculture. Targeted base editing of C-to-T per se and a phenotype verified in the generated mutants demonstrates the power of base editing in precise maize breeding.
文摘To confirm allelic relationship between Lr9 and the leaf rust resistance gene in KLM4-3B, genetics of resistance was studied using crosses (WL711 + Lr9) × WL711 and (WL711 + LrKLM4-3B) × WL711. The F2 populations in cross (WL711 + Lr9) × WL711 and (WL711 + LrKLM4-3B) × WL711 segregated in ratio of 3:1 for disease reaction at seedling stage against pathotype 77-5 of leaf rust. This suggests that rust resistance in these stocks are under the control of single dominant genes. Further, to study allelic relationship betweenLr 9 and LrKLM4-3B, F2 population of the cross (WL711 + LrKLM4-3B) × (WL711 + Lr9) was studied. A segregation ratio of 15:1 implies that the two genes Lr9 and LrKLM4-3B are non-allelic genes.
