Figure 3.

Comparison of the Ca²⁺ response in root tip cells expressing YC-Nano 65 or R-GECO1. Seedlings were exposed to a pulse of 10 µM NAA and medium containing either 1.5 mM calcium (CaCl₂) or 0-calcium. The schematic drawing on the right shows the root tip region examined. A, cpVenus/ECFP ratio at rest in root tip cells of YC-Nano 65 seedling in 1.5 mM calcium and 0-calcium. B, R-GECO1 fluorescence at rest in root tip cells of R-GECO1 seedlings in 1.5 mM calcium and 0-calcium. C, Normalized averaged YC-Nano 65 cpVenus/ECFP ratios (R) (ΔR/R₀ = (R−R₀)/R₀) over time in root tip cells in response to NAA as indicated by the black box on the x-axis. D, Normalized averaged R-GECO1 fluorescence (F) (ΔF/F₀ = (F−F₀)/F₀) over time in root tip cells in response to NAA as indicated by the black box on the x-axis. R₀ and F₀ are the prestimulus values of R and F, respectively. The shaded right-side arrow indicates the direction of the [Ca²⁺]_cyt increase. E, Maximal relative amplitude as ΔR_max/R₀ of cpVenus/ECFP ratio triggered by NAA in 1.5 mM calcium and 0-mM calcium conditions. F, Maximal relative amplitude as ΔF_max/F₀ of R-GECO1 fluorescence triggered by NAA in 1.5 mM calcium and 0-calcium conditions. G and H, Same experiments shown in (C and D) without normalization to the pre-stimulus situation. The shaded right-side arrow indicates the direction of the [Ca²⁺]_cyt increase. I, Maximal amplitude of cpVenus/ECFP ratio triggered by NAA application in 1.5 mM calcium and 0-calcium. J, Maximal amplitude of R-GECO1 fluorescence triggered by NAA application in 1.5 mM calcium and 0-calcium. Arbitrary units of fluorescence (AUF). n ≥ 6. Data were plotted as box-and-whisker plots using GraphPad, in which all the experimental points are plotted, and their distribution represented as a box that extends from the 25^th to 75^th percentiles. The line in the middle of the box is plotted at the median. P values were calculated with an unpaired Student’s t test. Error bars = sd. **P ≤ 0.005, ***P ≤ 0.0005 (t test).