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RootScan: Software for high-throughput analysis of root anatomical traits

Abstract

Background and aims

RootScan is a program for semi-automated image analysis of anatomical traits in root cross-sections.

Methods

RootScan uses pixel thresholds to separate the cross-section from its background and to divide it into tissue regions. Area measurements and object counts are performed within various regions of interest. A graphical user interface permits the user to see which regions are selected, to edit those selections, and to rate and comment on the data. The structure of the program allows for organized workflow and increased data collection efficiency.

Results

The program collects data on more than 20 variables per image including areas of the cross-section, stele, cortex, aerenchyma lacunae, xylem vessels, and counts of cortical cells and cell files. An increased rate of data collection allows collection of four times more variables in less time than is possible with current methods. Correlation analysis shows that RootScan data is equal or greater in accuracy than data collected with Photoshop.

Conclusions

Compared with currently available tools, this software offers considerable improvements in the amount and quality of data, ease of use, and time needed for data collection. RootScan permits phenotypic scoring of physiologically and agronomically important traits on a large number of genotypes.

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Abbreviations

GUI:

(graphical user interface)

RCA:

(root cortical aerenchyma)

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Acknowledgements

We thank Lauren Gelesh, Johanna Mirenda, Gina Riggio, Andy Evensen, Robert Snyder, and Chinmay Rao for technical assistance. United States Department of Agriculture, National Research Initiative provided funding for this research via grant 207-35100-18365 to JPL and KMB

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Authors

Corresponding author

Correspondence to Kathleen M. Brown.

Additional information

Responsible Editor: Matthias Wissuwa.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Online resource 1
figure 6

RootScan screenshots before and after lateral root selection in Zea mays. The user outlines the lateral root segment (a) and it is removed (b). (JPEG 114 kb)

Online resource 2
figure 7

Example of erroneous automatic cross-section selection. RootScan selected root hairs as part of this rice (Oryza sativa) root cross-section (red line). The user removed them by drawing a polygon that excluded the incorrect areas (blue line). Note that it is not necessary to draw the outline exactly, since the original selection will be used when it is inside the polygon. (JPEG 134 kb)

Online resource 3
figure 8

Rice root section images during aerenchyma selection and correction. The left image (a) shows that the program missed some aerenchyma lacunae, so the user manually selected the omitted objects by clicking with a crosshair tool. The right image (b) shows the aerenchyma lacunae in red after both RootScan and manual selection. (JPEG 215 kb)

Online resource 4
figure 9

RootScan incorrectly included part of the cortex of this field-grown maize root during the stele selection step (green line). The user redraws the stele boundary with a polygon tool (blue line). The error was caused by the high-contrast areas in the inner cortex due to fungal infection. (JPEG 124 kb)

Online resource 5
figure 10

The left image (a) shows a maize root section that has been sheared during sectioning. The cells and lacunae on the upper left are blurred. As a result, the lacunae in that region (image b, in red) are no longer completely visible and the area is reduced. The cells in that area are pixelated and cell counts would be too high. (JPEG 162 kb)

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High resolution image (TIFF 1876 kb)

High resolution image (TIFF 1876 kb)

High resolution image (TIFF 1876 kb)

High resolution image (TIFF 1876 kb)

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Burton, A.L., Williams, M., Lynch, J.P. et al. RootScan: Software for high-throughput analysis of root anatomical traits. Plant Soil 357, 189–203 (2012). https://doi.org/10.1007/s11104-012-1138-2

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  • DOI: https://doi.org/10.1007/s11104-012-1138-2

Keywords

  • Aerenchyma
  • Anatomy
  • Maize
  • Phenotyping
  • Trait
  • Root