Gfp filter

The coding sequence of the Ah24 gene, was fused with the 5′ region of green fluorescent protein (GFP) driven by the 35S CaMV promoter. The PCR product was amplified with specific primers listed in Table S5 of Supplementary Materials, and was subsequently cloned into the pDONR 221 plasmid and transferred to the pFAST-R06 plasmid using the Gateway technology (Invitrogen, Carlsbad, CA, USA). The binary vector was electroporated inro E. coli (DH5α strain) and A. tumefaciens (GV2260 strain). Arabidopsis plants (Col-0) were transformed with Agrobacterium using a modified floral dip method, as above. For GFP analysis, roots were observed under an inverted LSM510 confocal laser scanning microscope (Zeiss, Oberkochen, Germany). For visualization, seedlings were stained and mounted in 10 μg/ml propidium iodide (PI) solution (Sigma). GFP was excited with the 488 nm laser line of an argon laser, whereas PI was excited with the 514 nm laser line. The resulting images were acquired using the multi-channel mode. For GFP analysis of whole seedlings, a Lumar V.12 stereoscopic microscope with a GFP filter (Zeiss) was used.

YFP, GFP, and RFP fluorescence was observed with an Axio Imager.M2 microscope (Carl Zeiss Microscopy GmbH, Jena, Germany) with an epifluorescence HXP 120 illuminator (Zeiss) at 2–3 days post-infiltration (dpif). A GFP filter (Zeiss) with bandpass 470/40 nm for excitation and 525/50 nm for emission was used for GFP and YFP, and an RFP filter with bandpass 546/12 nm for excitation and 575–640 nm for emission was used for RFP. Images were acquired with an AxioCamMR3 controlled by ZEN 2012 blue edition software (Zeiss).
Confocal microscopy was carried out 3 dpif with a Leica TCS SP2 AOBS device using a 63× water immersion objective at the Institute of Biotechnology, University of Helsinki, as described [25 (link)], and the resulting images were analyzed using Leica LAS AF Lite.

The coding sequences of the AhERF and AhDof genes, were fused with the 5’ region of the green fluorescent protein (GFP) driven by the CaMV35 promoter. The constructions were subsequently employed to transform A. thaliana plants. The above procedures were performed as described previously [14 (link)]. For GFP analysis, roots were observed under an inverted LSM510 confocal laser scanning microscope (Zeiss, Oberkochen, Germany). For visualization, seedlings were stained and mounted in 10 μg/ ml propidium iodide (PI) solution (Sigma). The green fluorescent protein (GFP) was excited with the 488 nm laser line of an argon laser, whereas PI was excited with the 514 nm laser line. The resulting images were acquired using the multi-channel mode. For GFP analysis of whole seedlings, a Lumar V.12 stereoscopic microscope with a GFP filter (Carl Zeiss Microscopy GmbH, Jena, Germany) was used.