The operation of PEPCK increases light harvesting plasticity in C4 NAD-ME and NADP-ME photosynthetic subtypes: A theoretical study.

The repeated emergence of NADP-malic enzyme (ME), NAD-ME and phosphoenolpyruvate carboxykinase (PEPCK) subtypes of C4 photosynthesis are iconic examples of convergent evolution, which suggests that these biochemistries do not randomly assemble, but are instead specific adaptations resulting from unknown evolutionary drivers. Theoretical studies that are based on the classic biochemical understanding have repeatedly proposed light-use efficiency as a possible benefit of the PEPCK subtype. However, quantum yield measurements do not support this idea. We explore this inconsistency here via an analytical model that features explicit descriptions across a seamless gradient between C4 biochemistries to analyse light harvesting and dark photosynthetic metabolism. Our simulations show that the NADP-ME subtype, operated by the most productive crops, is the most efficient. The NAD-ME subtype has lower efficiency, but has greater light harvesting plasticity (the capacity to assimilate CO2 in the broadest combination of light intensity and spectral qualities). In both NADP-ME and NAD-ME backgrounds, increasing PEPCK activity corresponds to greater light harvesting plasticity but likely imposed a reduction in photosynthetic efficiency. We draw the first mechanistic links between light harvesting and C4 subtypes, providing the theoretical basis for future investigation.

Reduction of bundle sheath size boosts cyclic electron flow in C4 Setaria viridis acclimated to low light.

When C4 leaves are exposed to low light, the CO2 concentration in the bundle sheath (BS) cells decreases, causing an increase in photorespiration relative to assimilation, and a consequent reduction in biochemical efficiency. These effects can be mitigated by complex acclimation syndromes, which are of primary importance for crop productivity but are not well studied. We unveil an acclimation strategy involving the coordination of electron transport processes. First, we characterize the anatomy, gas exchange and electron transport of C4 Setaria viridis grown under low light. Through a purposely developed biochemical model, we resolve the photon fluxes and reaction rates to explain how the concerted acclimation strategies sustain photosynthetic efficiency. Our results show that a smaller BS in low-light-grown plants limited leakiness (the ratio of CO2 leak rate out of the BS over the rate of supply via C4 acid decarboxylation) but sacrificed light harvesting and ATP production. To counter ATP shortage and maintain high assimilation rates, plants facilitated light penetration through the mesophyll and upregulated cyclic electron flow in the BS. This shade tolerance mechanism, based on the optimization of light reactions, is possibly more efficient than the known mechanisms involving the rearrangement of carbon metabolism, and could potentially lead to innovative strategies for crop improvement.

C4 maize and sorghum are more sensitive to rapid dehydration than C3 wheat and sunflower.

The high productive potential, heat resilience, and greater water use efficiency of C4 over C3 plants attract considerable interest in the face of global warming and increasing population, but C4 plants are often sensitive to dehydration, questioning the feasibility of their wider adoption. To resolve the primary effect of dehydration from slower from secondary leaf responses originating within leaves to combat stress, we conducted an innovative dehydration experiment. Four crops grown in hydroponics were forced to a rapid yet controlled decrease in leaf water potential by progressively raising roots of out of the solution while measuring leaf gas exchange. We show that, under rapid dehydration, assimilation decreased more steeply in C4 maize and sorghum than in C3 wheat and sunflower. This reduction was due to a rise of nonstomatal limitation at triple the rate in maize and sorghum than in wheat and sunflower. Rapid reductions in assimilation were previously measured in numerous C4 species across both laboratory and natural conditions. Hence, we deduce that high sensitivity to rapid dehydration might stem from the disturbance of an intrinsic aspect of C4 bicellular photosynthesis. We posit that an obstruction to metabolite transport between mesophyll and bundle sheath cells could be the cause.

The slope of assimilation rate against stomatal conductance should not be used as a measure of water use efficiency or stomatal control over assimilation.

Quantifying water use efficiency, and the impact of stomata on CO2 uptake are pivotal in physiology and efforts to improve crop yields. Although tempting, relying on regression slopes from assimilation-stomatal conductance plots to estimate water use efficiency or stomatal control over assimilation is erroneous. Through numerical simulations, I substantiate this assertion. I propose the term 'instantaneous transpiration efficiency' for the assimilation-to-transpiration ratio to avoid confusion with 'intrinsic water use efficiency' which refers to the assimilation-to-stomatal conductance ratio, and recommend to compute both metrics for each gas exchange data point.

Upregulation of bundle sheath electron transport capacity under limiting light in C4 Setaria viridis.

C4 photosynthesis is a biochemical pathway that operates across mesophyll and bundle sheath (BS) cells to increase CO2 concentration at the site of CO2 fixation. C4 plants benefit from high irradiance but their efficiency decreases under shade, causing a loss of productivity in crop canopies. We investigated shade acclimation responses of Setaria viridis, a model monocot of NADP-dependent malic enzyme subtype, focussing on cell-specific electron transport capacity. Plants grown under low light (LL) maintained CO2 assimilation rates similar to high light plants but had an increased chlorophyll and light-harvesting-protein content, predominantly in BS cells. Photosystem II (PSII) protein abundance, oxygen-evolving activity and the PSII/PSI ratio were enhanced in LL BS cells, indicating a higher capacity for linear electron flow. Abundances of PSI, ATP synthase, Cytochrome b6 f and the chloroplast NAD(P)H dehydrogenase complex, which constitute the BS cyclic electron flow machinery, were also increased in LL plants. A decline in PEP carboxylase activity in mesophyll cells and a consequent shortage of reducing power in BS chloroplasts were associated with a more oxidised plastoquinone pool in LL plants and the formation of PSII - light-harvesting complex II supercomplexes with an increased oxygen evolution rate. Our results suggest that the supramolecular composition of PSII in BS cells is adjusted according to the redox state of the plastoquinone pool. This discovery contributes to the understanding of the acclimation of PSII activity in C4 plants and will support the development of strategies for crop improvement, including the engineering of C4 photosynthesis into C3 plants.

A leaf-level biochemical model simulating the introduction of C2 and C4 photosynthesis in C3 rice: gains, losses and metabolite fluxes.

This work aims at developing an adequate theoretical basis for comparing assimilation of the ancestral C3 pathway with CO2 concentrating mechanisms (CCM) that have evolved to reduce photorespiratory yield losses. We present a novel model for C3 , C2 , C2  + C4 and C4 photosynthesis simulating assimilatory metabolism, energetics and metabolite traffic at the leaf level. It integrates a mechanistic description of light reactions to simulate ATP and NADPH production, and a variable engagement of cyclic electron flow. The analytical solutions are compact and thus suitable for larger scale simulations. Inputs were derived with a comprehensive gas-exchange experiment. We show trade-offs in the operation of C4 that are in line with ecophysiological data. C4 has the potential to increase assimilation over C3 at high temperatures and light intensities, but this benefit is reversed under low temperatures and light. We apply the model to simulate the introduction of progressively complex levels of CCM into C3 rice, which feeds > 3.5 billion people. Increasing assimilation will require considerable modifications such as expressing the NAD(P)H Dehydrogenase-like complex and upregulating cyclic electron flow, enlarging the bundle sheath, and expressing suitable transporters to allow adequate metabolite traffic. The simpler C2 rice may be a desirable alternative.

A generalised dynamic model of leaf-level C3 photosynthesis combining light and dark reactions with stomatal behaviour.

Global food demand is rising, impelling us to develop strategies for improving the efficiency of photosynthesis. Classical photosynthesis models based on steady-state assumptions are inherently unsuitable for assessing biochemical and stomatal responses to rapid variations in environmental drivers. To identify strategies to increase photosynthetic efficiency, we need models that account for the timing of CO2 assimilation responses to dynamic environmental stimuli. Herein, I present a dynamic process-based photosynthetic model for C3 leaves. The model incorporates both light and dark reactions, coupled with a hydro-mechanical model of stomatal behaviour. The model achieved a stable and realistic rate of light-saturated CO2 assimilation and stomatal conductance. Additionally, it replicated complete typical assimilatory response curves (stepwise change in CO2 and light intensity at different oxygen levels) featuring both short lag times and full photosynthetic acclimation. The model also successfully replicated transient responses to changes in light intensity (light flecks), CO2 concentration, and atmospheric oxygen concentration. This dynamic model is suitable for detailed ecophysiological studies and has potential for superseding the long-dominant steady-state approach to photosynthesis modelling. The model runs as a stand-alone workbook in Microsoft® Excel® and is freely available to download along with a video tutorial.

Response of photosynthesis, growth and water relations of a savannah-adapted tree and grass grown across high to low CO2.

BACKGROUND AND AIMS: By the year 2100, atmospheric CO2 concentration ([CO2]a) could reach 800 ppm, having risen from ~200 ppm since the Neogene, beginning ~24 Myr ago. Changing [CO2]a affects plant carbon-water balance, with implications for growth, drought tolerance and vegetation shifts. The evolution of C4 photosynthesis improved plant hydraulic function under low [CO2]a and preluded the establishment of savannahs, characterized by rapid transitions between open C4-dominated grassland with scattered trees and closed forest. Understanding directional vegetation trends in response to environmental change will require modelling. But models are often parameterized with characteristics observed in plants under current climatic conditions, necessitating experimental quantification of the mechanistic underpinnings of plant acclimation to [CO2]a. METHODS: We measured growth, photosynthesis and plant-water relations, within wetting-drying cycles, of a C3 tree (Vachellia karroo, an acacia) and a C4 grass (Eragrostis curvula) grown at 200, 400 or 800 ppm [CO2]a. We investigated the mechanistic linkages between trait responses to [CO2]a under moderate soil drying, and photosynthetic characteristics. KEY RESULTS: For V. karroo, higher [CO2]a increased assimilation, foliar carbon:nitrogen, biomass and leaf starch, but decreased stomatal conductance and root starch. For Eragrostis, higher [CO2]a decreased C:N, did not affect assimilation, biomass or starch, and markedly decreased stomatal conductance. Together, this meant that C4 advantages in efficient water-use over the tree were maintained with rising [CO2]a. CONCLUSIONS: Acacia and Eragrostis acclimated differently to [CO2]a, with implications for their respective responses to water limitation and environmental change. Our findings question the carbon-centric focus on factors limiting assimilation with changing [CO2]a, how they are predicted and their role in determining productivity. We emphasize the continuing importance of water-conserving strategies in the assimilation response of savannah plants to rising [CO2]a.

Cell density and airspace patterning in the leaf can be manipulated to increase leaf photosynthetic capacity.

The pattern of cell division, growth and separation during leaf development determines the pattern and volume of airspace in a leaf. The resulting balance of cellular material and airspace is expected to significantly influence the primary function of the leaf, photosynthesis, and yet the manner and degree to which cell division patterns affect airspace networks and photosynthesis remains largely unexplored. In this paper we investigate the relationship of cell size and patterning, airspace and photosynthesis by promoting and repressing the expression of cell cycle genes in the leaf mesophyll. Using microCT imaging to quantify leaf cellular architecture and fluorescence/gas exchange analysis to measure leaf function, we show that increased cell density in the mesophyll of Arabidopsis can be used to increase leaf photosynthetic capacity. Our analysis suggests that this occurs both by increasing tissue density (decreasing the relative volume of airspace) and by altering the pattern of airspace distribution within the leaf. Our results indicate that cell division patterns influence the photosynthetic performance of a leaf, and that it is possible to engineer improved photosynthesis via this approach.

A Dynamic Hydro-Mechanical and Biochemical Model of Stomatal Conductance for C4 Photosynthesis.

C4 plants are major grain (maize [Zea mays] and sorghum [Sorghum bicolor]), sugar (sugarcane [Saccharum officinarum]), and biofuel (Miscanthus spp.) producers and contribute ∼20% to global productivity. Plants lose water through stomatal pores in order to acquire CO2 (assimilation [A]) and control their carbon-for-water balance by regulating stomatal conductance (gS). The ability to mechanistically predict gS and A in response to atmospheric CO2, water availability, and time is critical for simulating stomatal control of plant-atmospheric carbon and water exchange under current, past, or future environmental conditions. Yet, dynamic mechanistic models for gS are lacking, especially for C4 photosynthesis. We developed and coupled a hydromechanical model of stomatal behavior with a biochemical model of C4 photosynthesis, calibrated using gas-exchange measurements in maize, and extended the coupled model with time-explicit functions to predict dynamic responses. We demonstrated the wider applicability of the model with three additional C4 grass species in which interspecific differences in stomatal behavior could be accounted for by fitting a single parameter. The model accurately predicted steady-state responses of gS to light, atmospheric CO2 and oxygen, soil drying, and evaporative demand as well as dynamic responses to light intensity. Further analyses suggest that the effect of variable leaf hydraulic conductance is negligible. Based on the model, we derived a set of equations suitable for incorporation in land surface models. Our model illuminates the processes underpinning stomatal control in C4 plants and suggests that the hydraulic benefits associated with fast stomatal responses of C4 grasses may have supported the evolution of C4 photosynthesis.

A generalized stoichiometric model of C3, C2, C2+C4, and C4 photosynthetic metabolism.

The goal of suppressing photorespiration in crops to maximize assimilation and yield is stimulating considerable interest among researchers looking to bioengineer carbon-concentrating mechanisms into C3 plants. However, detailed quantification of the biochemical activities in the bundle sheath is lacking. This work presents a general stoichiometric model for C3, C2, C2+C4, and C4 assimilation (SMA) in which energetics, metabolite traffic, and the different decarboxylating enzymes (NAD-dependent malic enzyme, NADP-dependent malic enzyme, or phosphoenolpyruvate carboxykinase) are explicitly included. The SMA can be used to refine experimental data analysis or formulate hypothetical scenarios, and is coded in a freely available Microsoft Excel workbook. The theoretical underpinnings and general model behaviour are analysed with a range of simulations, including (i) an analysis of C3, C2, C2+C4, and C4 in operational conditions; (ii) manipulating photorespiration in a C3 plant; (iii) progressively upregulating a C2 shuttle in C3 photosynthesis; (iv) progressively upregulating a C4 cycle in C2 photosynthesis; and (v) manipulating processes that are hypothesized to respond to transient environmental inputs. Results quantify the functional trade-offs, such as the electron transport needed to meet ATP/NADPH demand, as well as metabolite traffic, inherent to different subtypes. The SMA refines our understanding of the stoichiometry of photosynthesis, which is of paramount importance for basic and applied research.

Systems biology and metabolic modelling unveils limitations to polyhydroxybutyrate accumulation in sugarcane leaves; lessons for C4 engineering.

In planta production of the bioplastic polyhydroxybutyrate (PHB) is one important way in which plant biotechnology can address environmental problems and emerging issues related to peak oil. However, high biomass C4 plants such as maize, switch grass and sugarcane develop adverse phenotypes including stunting, chlorosis and reduced biomass as PHB levels in leaves increase. In this study, we explore limitations to PHB accumulation in sugarcane chloroplasts using a systems biology approach, coupled with a metabolic model of C4 photosynthesis. Decreased assimilation was evident in high PHB-producing sugarcane plants, which also showed a dramatic decrease in sucrose and starch content of leaves. A subtle decrease in the C/N ratio was found which was not associated with a decrease in total protein content. An increase in amino acids used for nitrogen recapture was also observed. Based on the accumulation of substrates of ATP-dependent reactions, we hypothesized ATP starvation in bundle sheath chloroplasts. This was supported by mRNA differential expression patterns. The disruption in ATP supply in bundle sheath cells appears to be linked to the physical presence of the PHB polymer which may disrupt photosynthesis by scattering photosynthetically active radiation and/or physically disrupting thylakoid membranes.

Anatomical constraints to C4 evolution: light harvesting capacity in the bundle sheath.

In C4 photosynthesis CO2 assimilation and reduction are typically coordinated across mesophyll (M) and bundle sheath (BS) cells, respectively. This system consequently requires sufficient light to reach BS to generate enough ATP to allow ribulose-1,5-bisphosphate (RuBP) regeneration in BS. Leaf anatomy influences BS light penetration and therefore constrains C4 cycle functionality. Using an absorption scattering model (coded in Excel, and freely downloadable) we simulate light penetration profiles and rates of ATP production in BS across the C3 , C3 -C4 and C4 anatomical continua. We present a trade-off for light absorption between BS pigment concentration and space allocation. C3 BS anatomy limits light absorption and benefits little from high pigment concentrations. Unpigmented BS extensions increase BS light penetration. C4 and C3 -C4 anatomies have the potential to generate sufficient ATP in the BS, whereas typical C3 anatomy does not, except some C3 taxa closely related to C4 groups. Insufficient volume of BS, relative to M, will hamper a C4 cycle via insufficient BS light absorption. Thus, BS ATP production and RuBP regeneration, coupled with increased BS investments, allow greater operational plasticity. We propose that larger BS in C3 lineages may be co-opted for C3 -C4 and C4 biochemistry requirements.

Deriving C4 photosynthetic parameters from combined gas exchange and chlorophyll fluorescence using an Excel tool: theory and practice.

The higher photosynthetic potential of C4 plants has led to extensive research over the past 50 years, including C4 -dominated natural biomes, crops such as maize, or for evaluating the transfer of C4 traits into C3 lineages. Photosynthetic gas exchange can be measured in air or in a 2% Oxygen mixture using readily available commercial gas exchange and modulated PSII fluorescence systems. Interpretation of these data, however, requires an understanding (or the development) of various modelling approaches, which limit the use by non-specialists. In this paper we present an accessible summary of the theory behind the analysis and derivation of C4 photosynthetic parameters, and provide a freely available Excel Fitting Tool (EFT), making rigorous C4 data analysis accessible to a broader audience. Outputs include those defining C4 photochemical and biochemical efficiency, the rate of photorespiration, bundle sheath conductance to CO2 diffusion and the in vivo biochemical constants for PEP carboxylase. The EFT compares several methodological variants proposed by different investigators, allowing users to choose the level of complexity required to interpret data. We provide a complete analysis of gas exchange data on maize (as a model C4 organism and key global crop) to illustrate the approaches, their analysis and interpretation. © 2015 John Wiley & Sons Ltd.

An Excel tool for deriving key photosynthetic parameters from combined gas exchange and chlorophyll fluorescence: theory and practice.

Combined photosynthetic gas exchange and modulated fluorometres are widely used to evaluate physiological characteristics associated with phenotypic and genotypic variation, whether in response to genetic manipulation or resource limitation in natural vegetation or crops. After describing relatively simple experimental procedures, we present the theoretical background to the derivation of photosynthetic parameters, and provide a freely available Excel-based fitting tool (EFT) that will be of use to specialists and non-specialists alike. We use data acquired in concurrent variable fluorescence-gas exchange experiments, where A/Ci and light-response curves have been measured under ambient and low oxygen. From these data, the EFT derives light respiration, initial PSII (photosystem II) photochemical yield, initial quantum yield for CO2 fixation, fraction of incident light harvested by PSII, initial quantum yield for electron transport, electron transport rate, rate of photorespiration, stomatal limitation, Rubisco (ribulose 1·5-bisphosphate carboxylase/oxygenase) rate of carboxylation and oxygenation, Rubisco specificity factor, mesophyll conductance to CO2 diffusion, light and CO2 compensation point, Rubisco apparent Michaelis-Menten constant, and Rubisco CO2 -saturated carboxylation rate. As an example, a complete analysis of gas exchange data on tobacco plants is provided. We also discuss potential measurement problems and pitfalls, and suggest how such empirical data could subsequently be used to parameterize predictive photosynthetic models.

The operation of two decarboxylases, transamination, and partitioning of C4 metabolic processes between mesophyll and bundle sheath cells allows light capture to be balanced for the maize C4 pathway.

The C4 photosynthesis carbon-concentrating mechanism in maize (Zea mays) has two CO2 delivery pathways to the bundle sheath (BS; via malate or aspartate), and rates of phosphoglyceric acid reduction, starch synthesis, and phosphoenolpyruvate regeneration also vary between BS and mesophyll (M) cells. The theoretical partitioning of ATP supply between M and BS cells was derived for these metabolic activities from simulated profiles of light penetration across a leaf, with a potential 3-fold difference in the fraction of ATP produced in the BS relative to M (from 0.29 to 0.96). A steady-state metabolic model was tested using varying light quality to differentially stimulate M or BS photosystems. CO2 uptake, ATP production rate (JATP; derived with a low oxygen/chlorophyll fluorescence method), and carbon isotope discrimination were measured on plants under a low light intensity, which is considered to affect C4 operating efficiency. The light quality treatments did not change the empirical ATP cost of gross CO2 assimilation (JATP/GA). Using the metabolic model, measured JATP/GA was compared with the predicted ATP demand as metabolic functions were varied between M and BS. Transamination and the two decarboxylase systems (NADP-malic enzyme and phosphoenolpyruvate carboxykinase) were critical for matching ATP and reduced NADP demand in BS and M when light capture was varied under contrasting light qualities.

Acclimation to low light by C4 maize: implications for bundle sheath leakiness.

C4 plants have a biochemical carbon concentrating mechanism (CCM) that increases CO2 concentration around ribulose bisphosphate carboxylase oxygenase (Rubisco) in the bundle sheath (BS). Under limiting light, the activity of the CCM generally decreases, causing an increase in leakiness, (Φ), the ratio of CO2 retrodiffusing from the BS relative to C4 carboxylation processes. Maize plants were grown under high and low light regimes (respectively HL, 600 versus LL, 100 µE m(-2) s(-1) ). Short-term acclimation of Φ was compared from isotopic discrimination (Δ), gas exchange and photochemistry. Direct measurement of respiration in the light, and ATP production rate (JATP ), allowed us use a novel approach to derive Φ, compared with the conventional fitting of measured and predicted Δ. HL grown plants responded to decreasing light intensities with the well-documented increase in Φ. Conversely, LL plants showed a constant Φ, which has not been observed previously. We explain the pattern by two contrasting acclimation strategies: HL plants maintained a high CCM activity at LL, resulting in high CO2 overcycling and increased Φ; LL plants acclimated by down-regulating the CCM, effectively optimizing scarce ATP supply. This surprising plasticity may limit the impact of Φ-dependent carbon losses in leaves becoming shaded within developing canopies.

Acclimation of C4 metabolism to low light in mature maize leaves could limit energetic losses during progressive shading in a crop canopy.

C4 plants have a biochemical carbon-concentrating mechanism that increases CO2 concentration around Rubisco in the bundle sheath. Under low light, the activity of the carbon-concentrating mechanism generally decreases, associated with an increase in leakiness (ϕ), the ratio of CO2 retrodiffusing from the bundle sheath relative to C4 carboxylation. This increase in ϕ had been theoretically associated with a decrease in biochemical operating efficiency (expressed as ATP cost of gross assimilation, ATP/GA) under low light and, because a proportion of canopy photosynthesis is carried out by shaded leaves, potential productivity losses at field scale. Maize plants were grown under light regimes representing the cycle that leaves undergo in the canopy, whereby younger leaves initially developed under high light and were then re-acclimated to low light (600 to 100 µE·m(-2)·s(-1) photosynthetically active radiation) for 3 weeks. Following re-acclimation, leaves reduced rates of light-respiration and reached a status of lower ϕ, effectively optimizing the limited ATP resources available under low photosynthetically active radiation. Direct estimates of respiration in the light, and ATP production rate, allowed an empirical estimate of ATP production rate relative to gross assimilation to be derived. These values were compared to modelled ATP/GA which was predicted using leakiness as the sole proxy for ATP/GA, and, using a novel comprehensive biochemical model, showing that irrespective of whether leaves are acclimated to very low or high light intensity, the biochemical efficiency of the C4 cycle does not decrease at low photosynthetically active radiation.

A high throughput gas exchange screen for determining rates of photorespiration or regulation of C4 activity.

Large-scale research programmes seeking to characterize the C4 pathway have a requirement for a simple, high throughput screen that quantifies photorespiratory activity in C3 and C4 model systems. At present, approaches rely on model-fitting to assimilatory responses (A/C i curves, PSII quantum yield) or real-time carbon isotope discrimination, which are complicated and time-consuming. Here we present a method, and the associated theory, to determine the effectiveness of the C4 carboxylation, carbon concentration mechanism (CCM) by assessing the responsiveness of V O/V C, the ratio of RuBisCO oxygenase to carboxylase activity, upon transfer to low O2. This determination compares concurrent gas exchange and pulse-modulated chlorophyll fluorescence under ambient and low O2, using widely available equipment. Run time for the procedure can take as little as 6 minutes if plants are pre-adapted. The responsiveness of V O/V C is derived for typical C3 (tobacco, rice, wheat) and C4 (maize, Miscanthus, cleome) plants, and compared with full C3 and C4 model systems. We also undertake sensitivity analyses to determine the impact of R LIGHT (respiration in the light) and the effectiveness of the light saturating pulse used by fluorescence systems. The results show that the method can readily resolve variations in photorespiratory activity between C3 and C4 plants and could be used to rapidly screen large numbers of mutants or transformants in high throughput studies.

Computer reconstruction of plant growth and chlorophyll fluorescence emission in three spatial dimensions.

Plant leaves grow and change their orientation as well their emission of chlorophyll fluorescence in time. All these dynamic plant properties can be semi-automatically monitored by a 3D imaging system that generates plant models by the method of coded light illumination, fluorescence imaging and computer 3D reconstruction. Here, we describe the essentials of the method, as well as the system hardware. We show that the technique can reconstruct, with a high fidelity, the leaf size, the leaf angle and the plant height. The method fails with wilted plants when leaves overlap obscuring their true area. This effect, naturally, also interferes when the method is applied to measure plant growth under water stress. The method is, however, very potent in capturing the plant dynamics under mild stress and without stress. The 3D reconstruction is also highly effective in correcting geometrical factors that distort measurements of chlorophyll fluorescence emission of naturally positioned plant leaves.