It is increasingly important in life sciences that many cell-scale and tissue-scale measurements are quantified from confocal microscope images. However, extracting and analyzing large-scale confocal ...image data sets represents a major bottleneck for researchers. To aid this process, CellSeT software has been developed, which utilizes tissue-scale structure to help segment individual cells. We provide examples of how the CellSeT software can be used to quantify fluorescence of hormone-responsive nuclear reporters, determine membrane protein polarity, extract cell and tissue geometry for use in later modeling, and take many additional biologically relevant measures using an extensible plug-in toolset. Application of CellSeT promises to remove subjectivity from the resulting data sets and facilitate higher-throughput, quantitative approaches to plant cell research.
Seedling root traits impact plant establishment under challenging environments. Pearl millet is one of the most heat and drought tolerant cereal crops that provides a vital food source across the ...sub-Saharan Sahel region. Pearl millet's early root system features a single fast-growing primary root which we hypothesize is an adaptation to the Sahelian climate. Using crop modeling, we demonstrate that early drought stress is an important constraint in agrosystems in the Sahel where pearl millet was domesticated. Furthermore, we show that increased pearl millet primary root growth is correlated with increased early water stress tolerance in field conditions. Genetics including genome-wide association study and quantitative trait loci (QTL) approaches identify genomic regions controlling this key root trait. Combining gene expression data, re-sequencing and re-annotation of one of these genomic regions identified a glutaredoxin-encoding gene
as the candidate stress resilience root growth regulator. Functional characterization of its closest
homolog
revealed a novel role for this glutaredoxin (GRX) gene clade in regulating cell elongation. In summary, our study suggests a conserved function for GRX genes in conferring root cell elongation and enhancing resilience of pearl millet to its Sahelian environment.
Fluorescent reporters are valuable tools for plant science research, particularly as sensors to monitor biological signals and developmental processes. Such biosensors are particularly useful to ...monitor the spatial and temporal distribution of small signalling molecules such as phytohormones. Knowledge of perception and signalling pathways can be exploited to design biosensors for estimating intracellular abundance of hormones. However, the nonlinear relationship between target molecule and reporter necessitates the development of parameterised mathematical models to quantitatively relate sensor fluorescence to hormone abundance. In this opinion article, we will discuss the use of transcriptional reporters, sensor design strategy, and the importance of mathematical modelling approaches and technological advances in the development of new techniques to allow truly quantitative analyses of hormone regulated processes.
Studying the developmental genetics of plant organs requires following gene expression in specific tissues. To facilitate this, we have developed dual expression anatomy lines, which incorporate a ...red plasma membrane marker alongside a fluorescent reporter for a gene of interest in the same vector. Here, we adapted the GreenGate cloning vectors to create two destination vectors showing strong marking of cell membranes in either the whole root or specifically in the lateral roots. This system can also be used in both embryos and whole seedlings. As proof of concept, we follow both gene expression and anatomy in Arabidopsis (Arabidopsis thaliana) during lateral root organogenesis for a period of over 24 h. Coupled with the development of a flow cell and perfusion system, we follow changes in activity of the DII auxin sensor following application of auxin.
Root elongation and bending require the coordinated expansion of multiple cells of different types. These processes are regulated by the action of hormones that can target distinct cell layers. We ...use a mathematical model to characterise the influence of the biomechanical properties of individual cell walls on the properties of the whole tissue. Taking a simple constitutive model at the cell scale which characterises cell walls via yield and extensibility parameters, we derive the analogous tissue‐level model to describe elongation and bending. To accurately parameterise the model, we take detailed measurements of cell turgor, cell geometries and wall thicknesses. The model demonstrates how cell properties and shapes contribute to tissue‐level extensibility and yield. Exploiting the highly organised structure of the elongation zone (EZ) of the Arabidopsis root, we quantify the contributions of different cell layers, using the measured parameters. We show how distributions of material and geometric properties across the root cross‐section contribute to the generation of curvature, and relate the angle of a gravitropic bend to the magnitude and duration of asymmetric wall softening. We quantify the geometric factors which lead to the predominant contribution of the outer cell files in driving root elongation and bending.
Better understanding of root traits such as root angle and root gravitropism will be crucial for development of crops with improved resource use efficiency. This chapter describes a high-throughput, ...automated image analysis method to trace Arabidopsis (Arabidopsis thaliana) seedling roots grown on agar plates. The method combines a "particle-filtering algorithm with a graph-based method" to trace the center line of a root and can be adopted for the analysis of several root parameters such as length, curvature, and stimulus from original root traces.
Active Vision and Surface Reconstruction for 3D Plant Shoot Modelling Gibbs, Jonathon A.; Pound, Michael P.; French, Andrew P. ...
IEEE/ACM transactions on computational biology and bioinformatics,
2020-Nov.-Dec.-1, 2020 Nov-Dec, 2020-11-1, 20201101, Letnik:
17, Številka:
6
Journal Article
Recenzirano
Odprti dostop
Plant phenotyping is the quantitative description of a plant's physiological, biochemical, and anatomical status which can be used in trait selection and helps to provide mechanisms to link ...underlying genetics with yield. Here, an active vision- based pipeline is presented which aims to contribute to reducing the bottleneck associated with phenotyping of architectural traits. The pipeline provides a fully automated response to photometric data acquisition and the recovery of three-dimensional (3D) models of plants without the dependency of botanical expertise, whilst ensuring a non-intrusive and non-destructive approach. Access to complete and accurate 3D models of plants supports computation of a wide variety of structural measurements. An Active Vision Cell (AVC) consisting of a camera-mounted robot arm plus combined software interface and a novel surface reconstruction algorithm is proposed. This pipeline provides a robust, flexible, and accurate method for automating the 3D reconstruction of plants. The reconstruction algorithm can reduce noise and provides a promising and extendable framework for high throughput phenotyping, improving current state-of-the-art methods. Furthermore, the pipeline can be applied to any plant species or form due to the application of an active vision framework combined with the automatic selection of key parameters for surface reconstruction.
Root architecture impacts water and nutrient uptake efficiency. Identifying exactly which root architectural properties influence these agronomic traits can prove challenging. In this paper, ...approximately 300 wheat (Triticum aestivum) plants were divided into four groups using two binary classifications, high versus low nitrogen uptake efficiency (NUpE), and high versus low nitrate in the growth medium. The root system architecture for each wheat plant was captured using 16 quantitative variables. The multivariate analysis tool, linear discriminant analysis, was used to construct composite variables, each a linear combination of the original variables, such that the score of the plants on the new variables showed the maximum between-group variability. The results show that the distribution of root-system architecture traits differs between low- and high-NUpE plants and, less strongly, between low-NUpE plants grown on low versus high nitrate media.
Roots are highly responsive to environmental signals encountered in the rhizosphere, such as nutrients, mechanical resistance and gravity. As a result, root growth and development is very plastic. If ...this complex and vital process is to be understood, methods and tools are required to capture the dynamics of root responses. Tools are needed which are high-throughput, supporting large-scale experimental work, and provide accurate, high-resolution, quantitative data. We describe and demonstrate the efficacy of the high-throughput and high-resolution root imaging systems recently developed within the Centre for Plant Integrative Biology (CPIB). This toolset includes (i) robotic imaging hardware to generate time-lapse datasets from standard cameras under infrared illumination and (ii) automated image analysis methods and software to extract quantitative information about root growth and development both from these images and via high-resolution light microscopy. These methods are demonstrated using data gathered during an experimental study of the gravitropic response of Arabidopsis thaliana.
Summary
Image acquisition is an important step in the study of cytoskeleton organization. As visual interpretations and manual measurements of digital images are prone to errors and require a great ...amount of time, a freely available software package named MicroFilament Analyzer (MFA) was developed. The goal was to provide a tool that facilitates high‐throughput analysis to determine the orientation of filamentous structures on digital images in a more standardized, objective and repeatable way. Here, the rationale and applicability of the program is demonstrated by analyzing the microtubule patterns in epidermal cells of control and gravi‐stimulated Arabidopsis thaliana roots. Differential expansion of cells on either side of the root results in downward bending of the root tip. As cell expansion depends on the properties of the cell wall, this may imply a differential orientation of cellulose microfibrils. As cellulose deposition is orchestrated by cortical microtubules, the microtubule patterns were analyzed. The MFA program detects the filamentous structures on the image and identifies the main orientation(s) within individual cells. This revealed four distinguishable microtubule patterns in root epidermal cells. The analysis indicated that gravitropic stimulation and developmental age are both significant factors that determine microtubule orientation. Moreover, the data show that an altered microtubule pattern does not precede differential expansion. Other possible applications are also illustrated, including field emission scanning electron micrographs of cellulose microfibrils in plant cell walls and images of fluorescent actin.
PDF
