Plant-in-chip: Microfluidic system for studying root growth and pathogenic interactions in <i>Arabidopsis</i>

Parashar, Archana; Pandey, Santosh

doi:10.1063/1.3604788

Cited by 66 publications

(49 citation statements)

References 24 publications

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“…Over the past years, microfluidic devices have been developed for plant science to facilitate microscopic access to Arabidopsis roots (Meier et al, 2010;Grossmann et al, 2011;Parashar & Pandey, 2011;Busch et al, 2012;Jiang et al, 2014;Massalha et al, 2017), pollen tubes (Horade et al, 2013;Sanati Nezhad et al, 2013) or moss (Bascom et al, 2016). RootChips have been particularly useful for measuring cellular concentration changes of small molecules using genetically encoded nanosensors for nutrients (Grossmann et al, 2011;Lanquar et al, 2014), phytohormones (Jones et al, 2014) or the second messenger calcium (Denninger et al, 2014;Keinath et al, 2015).…”

Section: Researchmentioning

confidence: 99%

Dual‐flow‐RootChip reveals local adaptations of roots towards environmental asymmetry at the physiological and genetic levels

et al. 2017

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Roots grow in highly dynamic and heterogeneous environments. Biological activity as well as uneven nutrient availability or localized stress factors result in diverse microenvironments. Plants adapt their root morphology in response to changing environmental conditions, yet it remains largely unknown to what extent developmental adaptations are based on systemic or cell-autonomous responses. We present the dual-flow-RootChip, a microfluidic platform for asymmetric perfusion of Arabidopsis roots to investigate root-environment interactions under simulated environmental heterogeneity. Applications range from investigating physiology, root hair development and calcium signalling upon selective exposure to environmental stresses to tracing molecular uptake, performing selective drug treatments and localized inoculations with microbes. Using the dual-flow-RootChip, we revealed cell-autonomous adaption of root hair development under asymmetric phosphate (Pi) perfusion, with unexpected repression in root hair growth on the side exposed to low Pi and rapid tip-growth upregulation when Pi concentrations increased. The asymmetric root environment further resulted in an asymmetric gene expression of RSL4, a key transcriptional regulator of root hair growth. Our findings demonstrate that roots possess the capability to locally adapt to heterogeneous conditions in their environment at the physiological and transcriptional levels. Being able to generate asymmetric microenvironments for roots will help further elucidate decision-making processes in root-environment interactions.

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Section: Researchmentioning

confidence: 99%

Dual‐flow‐RootChip reveals local adaptations of roots towards environmental asymmetry at the physiological and genetic levels

et al. 2017

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“…Investigations of local root morphology responses in heterogenous settings with multiple, defined substrate sizes and chemistries will thus shed more light onto how plants respond to soil physiochemistry on a spatial and time scale. Multiple systems exist in which such experiments could be attempted, ranging from EcoFAB model systems to rhizotron designs (Grossmann et al, 2011; Parashar and Pandey, 2011; Rellán-Álvarez et al, 2015; Gao et al, 2018).…”

Section: Discussionmentioning

confidence: 99%

Root morphology and exudate availability is shaped by particle size and chemistry in Brachypodium distachyon

Sasse

Jordan

Raad

et al. 2019

Preprint

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Keywords Brachypodium distachyonParticle chemistry Particle size Pseudomonas fluorescens Rhizosphere Root exudation Root morphology Abstract Root morphology and exudation define a plants sphere of influence in soils, and are in turn shaped by the physiochemical characteristics of soil. We explored how particle size and chemistry of growth substrates affect root morphology and exudation of the model grass Brachypodium distachyon. Root fresh weight and root lengths were correlated with particle size, whereas root number and shoot weight remained constant. Mass spectrometry imaging suggested that both, root length and number shape root exudation. Exudate metabolite profiles detected with liquid chromatography / mass spectrometry were comparable for plants growing in glass beads or sand with various particles sizes, but distinct for plants growing in clay. However, when exudates of clay-grown plants were collected by removing the plants from the substrate, their exudate profile was similar to sand-or glass beads-grown plants. Clay particles sorbed 20% of compounds exuded by clay-grown plants, and 70% of compounds of a defined exudate medium. The sorbed compounds belonged to a range of chemical classes, among them nucleosides/nucleotides, organic acids, sugars, and amino acids. Some of the sorbed compounds could be de-sorbed by a rhizobacterium (Pseudomonas fluorescens WCS415), supporting its growth. We show that root morphology is affected by substrate size, and that root exudation in contrast is not affected by substrate size or chemistry. The availability of exuded compounds, however, depends on the substrate present. These findings further support the critical importance of the physiochemical properties of soils are crucial to consider when investigating plant morphology, exudation, and plantmicrobe interactions. Plant roots shape their environment in various ways, and are in turn shaped by physiochemical properties of the surrounding soil. Roots shape soil by dislocating particles, by polymer production, and by release of a wide variety of compounds (root exudation). Root exudates alter pH and the chemical composition around roots. Overall, root presence in soils results in formation of larger soil aggregates and increases water-holding capacity (Six et al., 2004). The plant-induced changes in chemistry can lead to weathering of minerals (Uroz et al., 2015), and alter the composition of microbial communities (Carson et al., 2007).Soils are often characterized by their particle size and mineralogy. Typical soil particles range from small (< 50 µm) to large (> 2 mm), determining physical parameters such as water binding capacity (Six et al., 2004; Six and Paustian, 2014; Rellán-Álvarez et al., 2016). Differently sized sandstones are associated with different microbial numbers, with small sandstones (2 mm) being more densely populated by microbes than larger rocks (Certini et al., 2004). Minerals differ in their structure (e.g. accessible surface), and in their surface charge, determining if and how they interact wi...

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“…Previously, seedlings were grown vertically on gelled media and transferred to a perfusion system immediately before the experiment, which allowed only measuring single roots at a time 8 . Microfluidic tools have been used for Arabidopsis, but on a low integration level 9 or without perfusion control 10 . The RootChip combines a high level of integration with the ability to automate experiments through precise flow guidance.…”

Section: Discussionmentioning

confidence: 99%

Time-lapse Fluorescence Imaging of Arabidopsis Root Growth with Rapid Manipulation of The Root Environment Using The RootChip

Großmann

Meier

Cartwright

et al. 2012

JoVE

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The root functions as the physical anchor of the plant and is the organ responsible for uptake of water and mineral nutrients such as nitrogen, phosphorus, sulfate and trace elements that plants acquire from the soil. If we want to develop sustainable approaches to producing high crop yield, we need to better understand how the root develops, takes up a wide spectrum of nutrients, and interacts with symbiotic and pathogenic organisms. To accomplish these goals, we need to be able to explore roots in microscopic detail over time periods ranging from minutes to days.We developed the RootChip, a polydimethylsiloxane (PDMS)-based microfluidic device, which allows us to grow and image roots from Arabidopsis seedlings while avoiding any physical stress to roots during preparation for imaging 1 (Figure 1). The device contains a bifurcated channel structure featuring micromechanical valves to guide the fluid flow from solution inlets to each of the eight observation chambers 2 . This perfusion system allows the root microenvironment to be controlled and modified with precision and speed. The volume of the chambers is approximately 400 nl, thus requiring only minimal amounts of test solution.Here we provide a detailed protocol for studying root biology on the RootChip using imaging-based approaches with real time resolution. Roots can be analyzed over several days using time lapse microscopy. Roots can be perfused with nutrient solutions or inhibitors, and up to eight seedlings can be analyzed in parallel. This system has the potential for a wide range of applications, including analysis of root growth in the presence or absence of chemicals, fluorescence-based analysis of gene expression, and the analysis of biosensors, e.g. FRET nanosensors 3 . Video LinkThe video component of this article can be found at https://www.jove.com/video/4290/ ProtocolNote: Perform all steps preparatory steps under sterile conditions. Preparation of Plastic Cones for Seed Germination1. Fill a 10 cm Petri dish with growth medium containing 1% agar to a thickness of 5 mm. We use a half strength modified Hoagland medium 4 , but medium composition should be chosen to suit individual experimental requirements. 2. While the medium is still liquid, use a multi-channel pipette to fill 10 μl pipette tips with 5 μl of medium from the Petri dish. 3. Store the filled tips in the pipette tip box until the medium is solid, then cut to 4 mm long plastic cones and place upright into a Petri dish containing solid growth medium. Seed Germination and Seedling Growth1. Surface sterilize seeds in 5% NaOCl for 5 min, wash three times with sterile water, then place a single seed on top of each of the mediumfilled cones. 2. Seal the dish with Micropore tape (3M) and store at 4 °C to synchronize germination. 3. After three days, transfer the plates to a growth cabinet to start germination. Our growth conditions are 23 °C at a 16h high light/8h dark cycle (Light intensity: 100 μE m -2 s -1).

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Plant-in-chip: Microfluidic system for studying root growth and pathogenic interactions in Arabidopsis

Cited by 66 publications

References 24 publications

Dual‐flow‐RootChip reveals local adaptations of roots towards environmental asymmetry at the physiological and genetic levels

Dual‐flow‐RootChip reveals local adaptations of roots towards environmental asymmetry at the physiological and genetic levels

Root morphology and exudate availability is shaped by particle size and chemistry in Brachypodium distachyon

Time-lapse Fluorescence Imaging of Arabidopsis Root Growth with Rapid Manipulation of The Root Environment Using The RootChip

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