X‐Ray Computed Tomography of Crop Plant Root Systems Grown in Soil

Stefan Mairhofer1, Tony Pridmore1, James Johnson1, Darren M. Wells1, Malcolm J. Bennett1, Sacha J. Mooney1, Craig J. Sturrock1

1 Centre for Plant Integrative Biology, School of Biosciences, University of Nottingham, Loughborough
Publication Name:  Current Protocols in Plant Biology
Unit Number:   
DOI:  10.1002/cppb.20049
Online Posting Date:  December, 2017
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Abstract

Non‐destructive methods to quantify the root system architecture of a plant grown in soil are essential to aid our understanding of the factors that impact plant root development in natural environments. With environmental change threatening our ability to sustain agricultural productivity for an expanding global population, the application of these methods has never before seen such an increase in demand. X‐ray computed tomography (CT) based phenotyping techniques permit the spatio‐temporal quantification of roots, helping to identify novel adaptive root architectural responses to abiotic and biotic factors. This protocol reports an integrated workflow from column preparation and plant growth to image and quantification of the root system using novel open source software applications, RooTrak and RooTh. © 2017 by John Wiley & Sons, Inc.

Keywords: image analysis; plant root phenotyping; root system architecture; soil; X‐ray computed tomography

     
 
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Table of Contents

  • Introduction
  • Basic Protocol 1: Soil Column Preparation
  • Basic Protocol 2: X‐Ray Computed Tomography (CT) Scanning
  • Basic Protocol 3: Using RooTrak for Root Material Recovery
  • Alternate Protocol 1: Using RooTrak Interactively (Edit)
  • Basic Protocol 4: Visualization and Quantification of Global Root System Traits
  • Basic Protocol 5: Quantification of Local Root System Traits and Standard RSML Output
  • Commentary
  • Literature Cited
  • Figures
  • Tables
     
 
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Materials

Basic Protocol 1: Soil Column Preparation

  Materials
  • Sugar beet seeds
  • Air dried, sieved (<2 mm) field soil
  • PVC or HDPP drain pipe
  • Hand saw or pipe cutter
  • Fine gauge mesh material
  • Scissors
  • Double‐sided adhesive tape
  • Heavy duty cotton cloth pressure sensitive adhesive tape (Gaffer tape)
  • 40‐liter plastic storage box
  • Measuring spoon
  • Balance

Basic Protocol 2: X‐Ray Computed Tomography (CT) Scanning

  Materials
  • Soil column (from protocol 1)
  • v|tome|x M 240 kV X‐ray CT system
  • Volume Graphics VGStudioMax v2.2
  • High performance workstation computer, recommended systems requirements: Intel Core i7 with 2.9 GHz and 32 to 64 GB of RAM (or higher)

Basic Protocol 3: Using RooTrak for Root Material Recovery

  Materials
  • RooTrak 0.3.9.1
  • High performance workstation computer, recommended systems requirements: Intel Core i7 with 2.9 GHz and 32‐64 GB of RAM (or higher)

Alternate Protocol 1: Using RooTrak Interactively (Edit)

  Materials
  • RooTrak 0.3.9.1
  • High performance workstation computer, recommended systems requirements: Intel Core i7 with 2.9 GHz and 32‐64 GB of RAM (or higher)

Basic Protocol 4: Visualization and Quantification of Global Root System Traits

  Materials
  • RooTrak 0.3.9.1
  • High performance workstation computer, recommended systems requirements: Intel Core i7 with 2.9 GHz and 32‐64 GB of RAM (or higher)

Basic Protocol 5: Quantification of Local Root System Traits and Standard RSML Output

  Materials
  • RooTh 0.5.88.52
  • High performance workstation computer, recommended systems requirements: Intel Core i7 with 2.9 GHz and 32‐64 GB of RAM (or higher)
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Figures

Videos

Literature Cited

  Gregory, P. J., Hutchison, D. J., Read, D. B., Jenneson, P. M., Gilboy, W. B., & Morton, E. J. (2003). Non‐invasive imaging of roots with high resolution X‐ray micro‐tomography. Plant and Soil, 255, 351–359. doi: 10.1023/a:1026179919689.
  Hargreaves, C. E., Gregory, P. J., & Bengough, A. G. (2009). Measuring root traits in barley (Hordeum vulgare ssp vulgare and ssp spontaneum) seedlings using gel chambers, soil sacs and X‐ray microtomography. Plant and Soil, 316, 285–297. doi: 10.1007/s11104‐008‐9780‐4.
  Heeraman, D. A., Hopmans, J. W., & Clausnitzer, V. (1997). Three dimensional imaging of plant roots in situ with x‐ray computed tomography. Plant and Soil, 189, 167–179. doi: 10.1023/a:1004258818538.
  Lafond, J. A., Han, L., & Dutilleul, P. (2015). Concepts and analyses in the CT scanning of root systems and leaf canopies: A timely summary. Frontiers in Plant Science, 6, 1111. doi: 10.3389/fpls.2015.01111.
  Lobet, G., Pound, M. P., Diener, J., Pradal, C., Draye, X., Godin, C., … Schnepf, A. (2015). Root system markup language: Toward a unified root architecture description language. Plant Physiology, 167, 617–627. doi: 10.1104/pp.114.253625.
  Mairhofer, S., Johnson, J., Sturrock, C. J., Bennett, M. J., Mooney, S. J., & Pridmore, T. P. (2015). Visual tracking for the recovery of multiple interacting plant root systems from X‐ray μCT images. Machine Vision and Applications, 27,721–734. . doi: 10.1007/s00138‐015‐0733‐7.
  Mairhofer, S., Zappala, S., Tracy, S. R., Sturrock, C., Bennett, M., Mooney, S. J., & Pridmore, T. (2012). RooTrak: Automated recovery of three‐dimensional plant root architecture in soil from x‐ray microcomputed tomography images using visual tracking. Plant Physiology, 158, 561–569. doi: 10.1104/pp.111.186221.
  Mairhofer, S., Zappala, S., Tracy, S., Sturrock, C., Bennett, M. J., Mooney, S. J., & Pridmore, T. P. (2013). Recovering complete plant root system architectures from soil via X‐ray mu‐Computed Tomography. Plant Methods, 9, 8. doi: 10.1186/1746‐4811‐9‐8 .
  Mooney, S., Pridmore, T., Helliwell, J., & Bennett, M. (2012). Developing X‐ray Computed Tomography to non‐invasively image 3‐D root systems architecture in soil. Plant and Soil, 352, 1–22. doi: 10.1007/s11104‐011‐1039‐9.
  Rogers, E. D., Monaenkova, D., Mijar, M., Nori, A., Goldman, D. I., & Benfey, P. N. (2016). X‐ray computed tomography reveals the response of root system architecture to soil texture. Plant Physiology, 171, 2028–2040. doi: 10.1104/pp.16.00397.
  Schmidt, S., Bengough, A. G., Gregory, P. J., Grinev, D. V., & Otten, W. (2012). Estimating root‐soil contact from 3D X‐ray microtomographs. European Journal of Soil Science, 63, 776–786. doi: 10.1111/j.1365‐2389.2012.01487.x.
  Sethian, J. A. (1999). Level set methods and fast marching methods: Evolving interfaces in computational geometry, fluid mechanics, computer vision, and materials science. Chapter 6: Basic Algorithms for Interface Evolution, Cambridge, United Kingdom: Cambridge University Press, 60–74.
  Sturrock, C. J., Woodhal, J., Brown, M., Walker, C., Mooney, S. J., & Ray, R. V. (2015). Effects of damping‐off caused by Rhizoctonia solani anastomosis group 2‐1 on roots of wheat and oil seed rape quantified using X‐ray Computed Tomography and real‐time PCR. Frontiers in Plant Science, 6, 461. doi: 10.3389/fpls.2015.00461.
  Tracy, S. R., Black, C. R., Roberts, J. A., Sturrock, C., Mairhofer, S., Craigon, J., & Mooney, S. J. (2012). Quantifying the impact of soil compaction on root system architecture in tomato (Solanum lycopersicum) by X‐ray micro‐computed tomography. Annals of Botany, 110, 511–519. doi: 10.1093/aob/mcs031.
  Zappala, S., Mairhofer, S., Tracy, S., Sturrock, C. J., Bennett, M., Pridmore, T., & Mooney, S. J. (2013). Quantifying the effect of soil moisture content on segmenting root system architecture in X‐ray computed tomography images. Plant and Soil, 370, 35–45. doi: 10.1007/s11104‐013‐1596‐1.
Internet Resources
  https://sourceforge.net/projects/rootrak
  Description and information for RooTrak, an open source software tool to explore and extract root system architecture.
  https://github.com/rootth/rooth
  Data repository for X‐ray CT image date for plant root systems.
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