ren activities  (Danaher Inc)

Journal: Horticulture Research

Article Title: LsAP2 regulates leaf morphology by inhibiting CIN-like TCP transcription factors and repressing LsKAN2 in lettuce

doi: 10.1038/s41438-021-00622-y

Figure Lengend Snippet: a Schematic representation of the LsAP2 protein structure. NLS nuclear localization sequence, EAR ethylene-responsive element binding factor-associated amphiphilic repression, AA amino acid. b Subcellular localization of LsAP2. c Schematic representation of the reporter and effector constructs used in the transcriptional activity assays. d Measurement of the relative firefly luciferase/Renilla luciferase (LUC/REN) ratio after transient coexpression of the reporter and effector constructs into tobacco leaves. The values are means ± SDs ( n = 6). The data were normalized to a value of 1 for the GAL4 DBD group. Different letters indicate significant differences determined by one-way ANOVA with Tukey’s post hoc test ( P < 0.05). e Mutation of the EAR motif (mEAR) in the LsAP2 protein sequence. f Yeast two-hybrid assays showing that LsAP2 interacts with LsTPL through the EAR motif. AD activation domain, BD binding domain. g Luciferase complementation imaging (LCI) assays showing the interaction between LsAP2 and N-LsTPL in tobacco leaves

Article Snippet: After 2 days of incubation, LUC and REN activities were measured using a SpectraMax ® i3x Multi-Mode detection platform (Molecular Devices, USA) with a Dual-Luciferase Reporter Assay Kit (Promega, USA).

Techniques: Sequencing, Binding Assay, Construct, Activity Assay, Luciferase, Mutagenesis, Activation Assay, Imaging

Journal: Horticulture Research

Article Title: LsAP2 regulates leaf morphology by inhibiting CIN-like TCP transcription factors and repressing LsKAN2 in lettuce

doi: 10.1038/s41438-021-00622-y

Figure Lengend Snippet: a Phylogenetic analysis of CIN-like TCP proteins. Ls Lactuca sativa , Am Antirrhinum majus , At Arabidopsis thaliana . The Antirrhinum CIN is indicated in red font. b Yeast two-hybrid assays showing the interactions between LsAP2 and CIN-like TCPs. We used 50 mM 3-amino-1,2,4-triazole (3-AT) for LsTCP3 and LsTCP4 and 0 mM 3-AT for the other LsTCPs. c LCI assays showing that LsAP2 interacts with LsTCP3 and LsTCP4. d Expression of CIN-like TCP genes in leaves. The values are means ± SDs ( n = 3). The expression data of LsTCP2 in leaves were normalized to 1. e Subcellular localization of LsTCP3 and LsTCP4 proteins. f Schematic representation of the reporter and effector constructs used in the transcriptional activity assays. g Measurement of the relative LUC/REN ratio after transient coexpression of the reporter and effector constructs in tobacco leaves. The values are means ± SDs ( n = 6). The data were normalized to a value of 1 for the GAL4 DBD group. Significant differences were determined by Student’s t test (** P < 0.01; *** P < 0.001)

Article Snippet: After 2 days of incubation, LUC and REN activities were measured using a SpectraMax ® i3x Multi-Mode detection platform (Molecular Devices, USA) with a Dual-Luciferase Reporter Assay Kit (Promega, USA).

Techniques: Expressing, Construct, Activity Assay

Journal: Horticulture Research

Article Title: LsAP2 regulates leaf morphology by inhibiting CIN-like TCP transcription factors and repressing LsKAN2 in lettuce

doi: 10.1038/s41438-021-00622-y

Figure Lengend Snippet: a Schematic representation of the LsKAN2 gene structure used for LsAP2 binding assays. b Yeast one-hybrid assays show that LsAP2 binds directly to the LsKAN2 promoter. Blue indicates an interaction. c Electrophoretic mobility shift assay (EMSA) showing that LsAP2 binds to the P3d fragment of the LsKAN2 promoter. MBP maltose-binding protein. The asterisk indicates nonspecific binding. d Schematic representation of the reporter and effector constructs used in the dual-luciferase reporter assays. e Measurement of the relative LUC/REN ratio after transient coexpression of pLsKAN2 : LUC with 35S:LsAP2 and 35S:LsTPL in tobacco leaves. The pGreenII 62-SK empty vector was used as the control. The values are means ± SDs ( n = 6). The data were normalized to a value of 1 for the control group. Significant differences were determined by Student’s t test (* P < 0.05; ** P < 0.01)

Article Snippet: After 2 days of incubation, LUC and REN activities were measured using a SpectraMax ® i3x Multi-Mode detection platform (Molecular Devices, USA) with a Dual-Luciferase Reporter Assay Kit (Promega, USA).

Techniques: Binding Assay, Electrophoretic Mobility Shift Assay, Construct, Luciferase, Plasmid Preparation