Quantitative 3-dimensional imaging of auxin and cytokinin levels in transgenic soybean and medicago truncatula roots via two-photon induced fluorescence imaging

Jon Fisher; Paul Gaillard; Ni Putu Dewi Nurmalasari; Carl Fellbaum; Sen Subramaniam; Steve Smith

doi:10.1117/12.2291288

23 February 2018 Quantitative 3-dimensional imaging of auxin and cytokinin levels in transgenic soybean and medicago truncatula roots via two-photon induced fluorescence imaging

Jon Fisher, Paul Gaillard, Ni Putu Dewi Nurmalasari, Carl Fellbaum, Sen Subramaniam, Steve Smith

Author Affiliations +

Proceedings Volume 10498, Multiphoton Microscopy in the Biomedical Sciences XVIII; 104982W (2018) https://doi.org/10.1117/12.2291288
Event: SPIE BiOS, 2018, San Francisco, California, United States

Abstract

Industrial nitrogen fertilizers account for nearly 50% of the fossil fuel costs in modern agriculture and contribute to soil and water pollution. Therefore, significant interest exists in understanding and characterizing the efficiency of nitrogen fixation, and the biochemical signaling pathways which orchestrate the plant-microbial symbiosis through which plants fix nitrogen. Legume plant species exhibit a particularly efficient nitrogen uptake mechanism, using root nodules which house nitrogen-fixing rhizobial bacteria. While nodule development has been widely studied, there remain significant gaps in understanding the regulatory hormones’ role in plant development. In this work, we produce 3-dimensional maps of auxin (AX) and cytokinin (CK) hormone concentrations within model plant root tips and nodules with respect to root architecture and cell type. Soybean and Medicago plants were transfected with a two-color fluorescent vector with AXsensitive green fluorescent protein (GFP) and CK-sensitive TdTomato (TdT). 3D images of soybean root nodules were captured using two-photon induced fluorescence microscopy. The resulting images were computationally analyzed using the localization code first developed by Weeks and later adapted by Kilfoil, and analyzed in the context of the root architecture. Statistical analysis of the resulting 3D hormone level maps reproduce-well the known roles of AX and CK in developing plant roots, and are the first quantitative description of these regulatory hormones tied to specific plant architecture. The analytical methods used, and the spatial distribution of these key regulatory hormones in plant roots, nodule primordia and root nodules, and their statistical interpretation are presented.

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Jon Fisher, Paul Gaillard, Ni Putu Dewi Nurmalasari, Carl Fellbaum, Sen Subramaniam, and Steve Smith "Quantitative 3-dimensional imaging of auxin and cytokinin levels in transgenic soybean and medicago truncatula roots via two-photon induced fluorescence imaging", Proc. SPIE 10498, Multiphoton Microscopy in the Biomedical Sciences XVIII, 104982W (23 February 2018); https://doi.org/10.1117/12.2291288

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KEYWORDS

Tissues

Luminescence

Nitrogen

Green fluorescent protein

Particles

Statistical analysis

Two photon imaging

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