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Regulatory pathway of stomatal development and target gene models. (a) Key proteins involved in the gene network regulating stomatal development in Arabidopsis (Pillitteri & Torii, ). The proteins targeted in this study are highlighted in green. (b) Gene models (not to scale) of the target genes STOMAGEN , EPF2 , YODA , and SPEECHLESS with their respective SolGenomcis ID and location within the Nicotiana benthamiana draft genome V1.0.1. The homeolog of each target gene is shown except for YODA , which has a single homeolog. The target sites of the sgRNAs are indicated with blue triangles in each target gene and its respective homeolog. SPEECHLESS A (Niben101Scf02511g04004.1) has a duplicated exon 1; hence, sgRNA1 targets this homolog at two sites. (c) TRV1 and <t>TRV2</t> vector component and design.
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Regulatory pathway of stomatal development and target gene models. (a) Key proteins involved in the gene network regulating stomatal development in Arabidopsis (Pillitteri & Torii, ). The proteins targeted in this study are highlighted in green. (b) Gene models (not to scale) of the target genes STOMAGEN , EPF2 , YODA , and SPEECHLESS with their respective SolGenomcis ID and location within the Nicotiana benthamiana draft genome V1.0.1. The homeolog of each target gene is shown except for YODA , which has a single homeolog. The target sites of the sgRNAs are indicated with blue triangles in each target gene and its respective homeolog. SPEECHLESS A (Niben101Scf02511g04004.1) has a duplicated exon 1; hence, sgRNA1 targets this homolog at two sites. (c) TRV1 and <t>TRV2</t> vector component and design.
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Regulatory pathway of stomatal development and target gene models. (a) Key proteins involved in the gene network regulating stomatal development in Arabidopsis (Pillitteri & Torii, ). The proteins targeted in this study are highlighted in green. (b) Gene models (not to scale) of the target genes STOMAGEN , EPF2 , YODA , and SPEECHLESS with their respective SolGenomcis ID and location within the Nicotiana benthamiana draft genome V1.0.1. The homeolog of each target gene is shown except for YODA , which has a single homeolog. The target sites of the sgRNAs are indicated with blue triangles in each target gene and its respective homeolog. SPEECHLESS A (Niben101Scf02511g04004.1) has a duplicated exon 1; hence, sgRNA1 targets this homolog at two sites. (c) TRV1 and <t>TRV2</t> vector component and design.
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Regulatory pathway of stomatal development and target gene models. (a) Key proteins involved in the gene network regulating stomatal development in Arabidopsis (Pillitteri & Torii, ). The proteins targeted in this study are highlighted in green. (b) Gene models (not to scale) of the target genes STOMAGEN , EPF2 , YODA , and SPEECHLESS with their respective SolGenomcis ID and location within the Nicotiana benthamiana draft genome V1.0.1. The homeolog of each target gene is shown except for YODA , which has a single homeolog. The target sites of the sgRNAs are indicated with blue triangles in each target gene and its respective homeolog. SPEECHLESS A (Niben101Scf02511g04004.1) has a duplicated exon 1; hence, sgRNA1 targets this homolog at two sites. (c) TRV1 and <t>TRV2</t> vector component and design.
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Regulatory pathway of stomatal development and target gene models. (a) Key proteins involved in the gene network regulating stomatal development in Arabidopsis (Pillitteri & Torii, ). The proteins targeted in this study are highlighted in green. (b) Gene models (not to scale) of the target genes STOMAGEN , EPF2 , YODA , and SPEECHLESS with their respective SolGenomcis ID and location within the Nicotiana benthamiana draft genome V1.0.1. The homeolog of each target gene is shown except for YODA , which has a single homeolog. The target sites of the sgRNAs are indicated with blue triangles in each target gene and its respective homeolog. SPEECHLESS A (Niben101Scf02511g04004.1) has a duplicated exon 1; hence, sgRNA1 targets this homolog at two sites. (c) TRV1 and <t>TRV2</t> vector component and design.
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Regulatory pathway of stomatal development and target gene models. (a) Key proteins involved in the gene network regulating stomatal development in Arabidopsis (Pillitteri & Torii, ). The proteins targeted in this study are highlighted in green. (b) Gene models (not to scale) of the target genes STOMAGEN , EPF2 , YODA , and SPEECHLESS with their respective SolGenomcis ID and location within the Nicotiana benthamiana draft genome V1.0.1. The homeolog of each target gene is shown except for YODA , which has a single homeolog. The target sites of the sgRNAs are indicated with blue triangles in each target gene and its respective homeolog. SPEECHLESS A (Niben101Scf02511g04004.1) has a duplicated exon 1; hence, sgRNA1 targets this homolog at two sites. (c) TRV1 and <t>TRV2</t> vector component and design.
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Regulatory pathway of stomatal development and target gene models. (a) Key proteins involved in the gene network regulating stomatal development in Arabidopsis (Pillitteri & Torii, ). The proteins targeted in this study are highlighted in green. (b) Gene models (not to scale) of the target genes STOMAGEN , EPF2 , YODA , and SPEECHLESS with their respective SolGenomcis ID and location within the Nicotiana benthamiana draft genome V1.0.1. The homeolog of each target gene is shown except for YODA , which has a single homeolog. The target sites of the sgRNAs are indicated with blue triangles in each target gene and its respective homeolog. SPEECHLESS A (Niben101Scf02511g04004.1) has a duplicated exon 1; hence, sgRNA1 targets this homolog at two sites. (c) TRV1 and TRV2 vector component and design.

Journal: The Plant Journal

Article Title: Virus‐induced gene editing of stomatal regulators in Nicotiana benthamiana enables rapid functional genomics

doi: 10.1111/tpj.70805

Figure Lengend Snippet: Regulatory pathway of stomatal development and target gene models. (a) Key proteins involved in the gene network regulating stomatal development in Arabidopsis (Pillitteri & Torii, ). The proteins targeted in this study are highlighted in green. (b) Gene models (not to scale) of the target genes STOMAGEN , EPF2 , YODA , and SPEECHLESS with their respective SolGenomcis ID and location within the Nicotiana benthamiana draft genome V1.0.1. The homeolog of each target gene is shown except for YODA , which has a single homeolog. The target sites of the sgRNAs are indicated with blue triangles in each target gene and its respective homeolog. SPEECHLESS A (Niben101Scf02511g04004.1) has a duplicated exon 1; hence, sgRNA1 targets this homolog at two sites. (c) TRV1 and TRV2 vector component and design.

Article Snippet: TRV‐mobile sgRNA vectors were assembled by amplifying the sgRNA scaffold and truncated FT sequence from TRV2 T‐DNA vector pEE515 (Addgene plasmid number 149288).

Techniques: Plasmid Preparation

Infection and somatic editing with TRV‐AmCyan vectors. (a) Vector design of TRV1 and TRV2‐AmCyan with the mobile sgRNA cloned downstream of AmCyan. (b) AmCyan signals in new leaves of infected plant demonstrating systemic movement of TRV2 (left), representative images of adaxial epidermal peels with varying degrees of stomatal density (right). (c) Somatic editing frequency at four target loci following delivery of mobile sgRNAs using TRV2 vectors (turquoise) or TRV2‐AmCyan–mobile sgRNA vectors (purple). Bars represent mean editing frequency (%) and error bars indicate ± SD. (d) Stomatal density in AmCyan‐positive M 0 plants ( n = 2) across control (AmCyan) and sgRNA‐treated groups. Bars represent mean stomatal density (mm −2 ), and error bars indicate ± SD. Asterisks denote statistically significant differences ( P < 0.05) compared with the control.

Journal: The Plant Journal

Article Title: Virus‐induced gene editing of stomatal regulators in Nicotiana benthamiana enables rapid functional genomics

doi: 10.1111/tpj.70805

Figure Lengend Snippet: Infection and somatic editing with TRV‐AmCyan vectors. (a) Vector design of TRV1 and TRV2‐AmCyan with the mobile sgRNA cloned downstream of AmCyan. (b) AmCyan signals in new leaves of infected plant demonstrating systemic movement of TRV2 (left), representative images of adaxial epidermal peels with varying degrees of stomatal density (right). (c) Somatic editing frequency at four target loci following delivery of mobile sgRNAs using TRV2 vectors (turquoise) or TRV2‐AmCyan–mobile sgRNA vectors (purple). Bars represent mean editing frequency (%) and error bars indicate ± SD. (d) Stomatal density in AmCyan‐positive M 0 plants ( n = 2) across control (AmCyan) and sgRNA‐treated groups. Bars represent mean stomatal density (mm −2 ), and error bars indicate ± SD. Asterisks denote statistically significant differences ( P < 0.05) compared with the control.

Article Snippet: TRV‐mobile sgRNA vectors were assembled by amplifying the sgRNA scaffold and truncated FT sequence from TRV2 T‐DNA vector pEE515 (Addgene plasmid number 149288).

Techniques: Infection, Plasmid Preparation, Clone Assay, Control