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Adaptation of high-efficiency CRISPR/Cas9-based multiplex genome editing system in white lupin by using endogenous promoters.
Zhu, Xiaoqi; Xu, Weifeng; Liu, Bowen; Zhan, Yujie; Xia, Tianyu.
Afiliação
  • Zhu X; Joint International Research Laboratory of Water and Nutrient in Crop and College of Resource and Environment, Center for Plant Water-use and Nutrition Regulation and College of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou, China.
  • Xu W; Joint International Research Laboratory of Water and Nutrient in Crop and College of Resource and Environment, Center for Plant Water-use and Nutrition Regulation and College of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou, China.
  • Liu B; Joint International Research Laboratory of Water and Nutrient in Crop and College of Resource and Environment, Center for Plant Water-use and Nutrition Regulation and College of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou, China.
  • Zhan Y; Joint International Research Laboratory of Water and Nutrient in Crop and College of Resource and Environment, Center for Plant Water-use and Nutrition Regulation and College of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou, China.
  • Xia T; Joint International Research Laboratory of Water and Nutrient in Crop and College of Resource and Environment, Center for Plant Water-use and Nutrition Regulation and College of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou, China.
Physiol Plant ; 175(4): e13976, 2023.
Article em En | MEDLINE | ID: mdl-37616014
ABSTRACT
White lupin (Lupinus albus L.) is an important crop with high phosphorus (P) use efficiency; however, technologies for its functional genomic and molecular analyses are limited. Cluster regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 (Cas9) (CRISPR/Cas9) system has been applied to gene editing and function genomics in many crops, but its application in white lupin has not been well documented. Here, we adapted the CRISPR/Cas9-based multiplex genome editing system by using the native U3/U6 and ubiquitin (UBQ) promoters to drive sgRNAs and Cas9. Two target sites (T1 and T2) within the Lalb_Chr05g0223881 gene, encoding a putative trehalase, were selected to validate its efficacy in white lupin based on the Agrobacterium rhizogenes-mediated transformation. We found that the T0 hairy roots were efficiently mutated at T1 and T2 with a frequency of 6.25%-35% and 50%-92.31%, respectively. The mutation types include nucleotide insertion, deletion, substitution, and complicated variant. Simultaneous mutations of the two targets were also observed with a range of 6.25%-35%. The combination of LaU6.6 promoter for sgRNA and LaUBQ12 promoter for Cas9 generated the highest frequency of homozygous/biallelic mutations at 38.46%. In addition, the target-sgRNA sequence also contributes to the editing efficiency of the CRISPR/Cas9 system in white lupin. In conclusion, our results expand the toolbox of the CRISPR/Cas9 system and benefit the basic research in white lupin.
Assuntos

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Lupinus / Edição de Genes Idioma: En Revista: Physiol Plant Ano de publicação: 2023 Tipo de documento: Article País de afiliação: China

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Lupinus / Edição de Genes Idioma: En Revista: Physiol Plant Ano de publicação: 2023 Tipo de documento: Article País de afiliação: China