植物科学

Researchers face a number of challenges in the construction of soil columns which can affect the outcome of their experiments. The use of intact soil cores closely mimics actual field conditions. However, the excavation of large intact soil cores is a time-consuming, labor-intensive process and may lead to soil compaction that would influence the solute transport behavior of the soil column. Repacked soil columns are used as an option to circumvent these challenges of intact soil cores. However, repacked soil columns also have their limitations and introduce other challenges. Here, we present a step by step procedure for the design of repacked soil columns to achieve a realistic bulk density, prevent preferential flow paths, and ensure hydraulic connectivity between soil layers. This protocol will be beneficial to Soil Scientists, Hydrologists and other Environmental Scientists utilizing repacked soil columns.

Field studies that simulate the effects of climate change are important for a predictive understanding of ecosystem responses to a changing environment. Among many concerns, regional warming can result in advanced timing of spring snowmelt in snowpack dependent ecosystems, which could lead to longer snow-free periods and drier summer soils. Past studies investigating these impacts of climate change have manipulated snowmelt with a variety of techniques that include manual snowpack alteration with a shovel, infrared radiation, black sand and fabric covers. Within these studies however, sufficient documentation of methods is limited, which can make experimental reproduction difficult. Here, we outline a detailed plot-scale protocol that utilizes a permeable black geotextile fabric deployed on top of an isothermal spring snowpack to induce advanced snowmelt. The method offers a reliable and cost-effective approach to induce snowmelt by passively increasing solar radiation absorption at the snow surface. In addition, control configurations with no snowpack manipulation are paired adjacent to the induced snowmelt plot for experimental comparison. Past and ongoing deployments in Colorado subalpine ecosystems indicate that this approach can accelerate snowmelt by 14-23 days, effectively mimicking snowmelt timing at lower elevations. This protocol can be applied to a variety of studies to understand the hydrological, ecological, and geochemical impacts of regional warming in snowpack dependent ecosystems.

Acclimation of leaf traits to fluctuating environments is a key mechanism to maximize fitness. One of the most important strategies in acclimation to changing light is to maintain efficient utilization of nitrogen in the photosynthetic apparatus by continuous modifications of between-leaf distribution along the canopy depth and within-leaf partitioning between photosynthetic functions according to local light availability. Between-leaf nitrogen distribution has been intensively studied over the last three decades, where proportional coordination between nitrogen concentration and light gradient was considered optimal in terms of maximizing canopy photosynthesis, without taking other canopy structural and physiological factors into account. We proposed a mechanistic model of protein turnover dynamics in different photosynthetic functions, which can be parameterized using leaves grown under different levels of constant light. By integrating this dynamic model into a multi-layer canopy model, constructed using data collected from a greenhouse experiment, it allowed us to test in silico the degree of optimality in photosynthetic nitrogen use for maximizing canopy carbon assimilation under given light environments.

The plant cell wall is a complicated network that is mainly constituted of polysaccharides, such as cellulose, hemicellulose and pectin. Many noncellulosic polysaccharides are further acetylated, which confers these polymers flexible physicochemical properties. Due to the significance of cell wall in plant growth and development, the analytic platform has been the focus for a long time. Here, we use internodes/culms, an important organ to provide mechanical support for rice plants, as an experimental sample to explore the method for cell wall composition analysis. The method includes preparation of cell wall residues, sequential extraction of polysaccharides, and measurement of cellulose. The procedure for acetate examination is also described. This method is applicable to determine the composition of individual cell wall polymers and the modifier acetates, and is suitable to identify cell wall relevant mutants based on the advantages in high throughput, precision and repeatability.

Biogenic volatile compounds (VCs) mediate various types of crucial intra- and inter-species interactions in plants, animals, and microorganisms owing to their ability to travel through air, liquid, and porous soils. To study how VCs produced by Verticillium dahliae, a soilborne fungal pathogen, affect plant growth and development, we slightly modified a method previously used to study the effect of bacterial VCs on plant growth. The method involves culturing microbial cells and plants in I plate to allow only VC-mediated interaction. The modified protocol is simple to set up and produces reproducible results, facilitating studies on this poorly explored form of plant-fungal interactions. We also optimized conditions for extracting and identifying fungal VCs using solid phase microextraction (SPME) coupled to gas chromatography-mass spectrometry (GC-MS).

High-throughput phenotyping of plant traits is a powerful tool to further our understanding of plant growth and its underlying physiological, molecular, and genetic determinisms. This protocol describes the methodology of a standard phenotyping experiment in PHENOPSIS automated platform, which was engineered in INRA-LEPSE (https://www6.montpellier.inra.fr/lepse) and custom-made by Optimalog company. The seminal method was published by Granier et al. (2006). The platform is used to explore and test various ecophysiological hypotheses (Tisné et al., 2010; Baerenfaller et al., 2012; Vile et al., 2012; Bac-Molenaar et al., 2015; Rymaszewski et al., 2017). Here, the focus concerns the preparation and management of experiments, as well as measurements of growth-related traits (e.g., projected rosette area, total leaf area and growth rate), water status-related traits (e.g., leaf dry matter content and relative water content), and plant architecture-related traits (e.g., stomatal density and index and lamina/petiole ratio). Briefly, a completely randomized (block) design is set up in the growth chamber. Next, the substrate is prepared, its initial water content is measured and pots are filled. Seeds are sown onto the soil surface and germinated prior to the experiment. After germination, soil watering and image (visible, infra-red, fluorescence) acquisition are planned by the user and performed by the automaton. Destructive measurements may be performed during the experiment. Data extraction from images and estimation of growth-related trait values involves semi-automated procedures and statistical processing.

We have proposed and tested a method for characterization of the signal sequences and determinations of target protein localization in a plant cell. This method, called the AgI-PrI, implies extraction of protoplasts from plant tissues after agroinfiltration. The suggested approach combines the advantages of two widely used methods for transient gene expression in plants–agroinfiltration and transfection of isolated protoplasts. The AgI-PrI technic can be applied to other plant species.

The rapid auxin-triggered growth of the Arabidopsis hypocotyls involves the nuclear TIR1/AFB-Aux/IAA signaling and is accompanied by acidification of the apoplast and cell walls (Fendrych et al., 2016). Here, we describe in detail the method for analysis of the elongation and the TIR1/AFB-Aux/IAA-dependent auxin response in hypocotyl segments as well as the determination of relative values of the cell wall pH.

It is well-established that plants are able to acclimate to temperatures above or below the optimal temperature for their growth. Here, we provide protocols for assays that can be used quantitatively or qualitatively to assess the relative ability of plants to acquire tolerance to high temperature stress. The hypocotyl elongation assay described was developed to screen for mutants defective in the acquisition of tolerance to extreme temperature stress, and other assays were developed to further characterize mutant and transgenic plants for heat tolerance of other processes or at other growth stages. Although the protocols provide details for application to Arabidopsis thaliana, the same basic methods can be adopted to assay heat tolerance in other plant species.

Phenotyping the dynamics of root responses to environmental cues is necessary to understand plant acclimation to their environment. Continuous monitoring of root growth is challenging because roots normally grow belowground and are very sensitive to their growth environment. This protocol combines infrared imaging with hydroponic cultivation for kinematic analyses. It allows continuous imaging at fine spatiotemporal resolution and disturbs roots minimally. Examples are provided of how the procedure and materials can be adapted for 3D monitoring and of how environmental stress may be manipulated for experimental purposes.