Planta

, Volume 234, Issue 4, pp 769–784 | Cite as

Automated motion estimation of root responses to sucrose in two Arabidopsis thaliana genotypes using confocal microscopy

  • Nathalie Wuyts
  • A. Glyn Bengough
  • Timothy J. Roberts
  • Chengjin Du
  • M. Fraser Bransby
  • Stephen J. McKenna
  • Tracy A. Valentine
Original Article

Abstract

Root growth is a highly dynamic process influenced by genetic background and environment. This paper reports the development of R scripts that enable root growth kinematic analysis that complements a new motion analysis tool: PlantVis. Root growth of Arabidopsis thaliana expressing a plasma membrane targeted GFP (C24 and Columbia 35S:LTI6b-EGFP) was imaged using time-lapse confocal laser scanning microscopy. Displacement of individual pixels in the time-lapse sequences was estimated automatically by PlantVis, producing dense motion vector fields. R scripts were developed to extract kinematic growth parameters and report displacement to ±0.1 pixel. In contrast to other currently available tools, Plantvis-R delivered root velocity profiles without interpolation or averaging across the root surface and also estimated the uncertainty associated with tracking each pixel. The PlantVis-R analysis tool has a range of potential applications in root physiology and gene expression studies, including linking motion to specific cell boundaries and analysis of curvature. The potential for quantifying genotype × environment interactions was examined by applying PlantVis-R in a kinematic analysis of root growth of C24 and Columbia, under contrasting carbon supply. Large genotype-dependent effects of sucrose were recorded. C24 exhibited negligible differences in elongation zone length and elongation rate but doubled the density of lateral roots in the presence of sucrose. Columbia, in contrast, increased its elongation zone length and doubled its elongation rate and the density of lateral roots.

Keywords

Cell expansion Confocal laser scanning microscopy Digital image analysis Motion estimation Root meristem Sucrose 

Abbreviations

CLSM

Confocal laser scanning microscopy

DAG

Days after germination

DVF

Displacement vector field

EGR

Elemental growth rate

PIV

Particle image velocimetry

Supplementary material

425_2011_1435_MOESM1_ESM.tif (1.8 mb)
Fig. S1 Motion estimation in a time-lapse image sequence of an A. thaliana primary root. PlantVis was applied on a CLSM image sequence (1 min between frames), acquired using transmission settings, of an A. thaliana C24 LTI6b-EGFP primary root (10 DAG) at 10× magnification. Results were imported into R for analysis and visualisation. a Confocal image. b Absolute velocity (μm min−1) (TIFF 1,869 kb)
425_2011_1435_MOESM2_ESM.mpg (186 kb)
Fig. S2 Video of a time-lapse CLSM image sequence (1 min between frames) of an A. thaliana LTI6b-EGFP primary root (8 DAG) at 10× magnification. a C24. b Col (MPG 186 kb)
425_2011_1435_MOESM3_ESM.mpg (222 kb)
Supplementary material 3 (MPG 222 kb)
425_2011_1435_MOESM4_ESM.tif (2.4 mb)
Fig. S3 Scatterplots of growth parameters for individual A. thaliana primary roots. PlantVis was applied on CLSM image sequences (1 min between frames) of A. thaliana C24 LTI6b-EGFP primary roots (5, 8 and 11 DAG) at 10× magnification. Growth parameters are plotted against longitudinal velocity for three individual roots (black, dark grey, light grey) at 5, 8 and 11 DAG, including root length (a), growth zone length (b), elongation zone length (c), division zone length (d), maximum elemental growth rate (e), and position of maximum elemental growth rate (f) along the root axis (as distance from the quiescent centre) (TIFF 2,472 kb)
425_2011_1435_MOESM5_ESM.rtf (42 kb)
Supplementary Table S1 (RTF 42 kb)
425_2011_1435_MOESM6_ESM.doc (36 kb)
Supplementary Table S2 (DOC 36 kb)

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Copyright information

© Springer-Verlag 2011

Authors and Affiliations

  • Nathalie Wuyts
    • 1
    • 4
  • A. Glyn Bengough
    • 1
    • 3
  • Timothy J. Roberts
    • 2
  • Chengjin Du
    • 2
  • M. Fraser Bransby
    • 3
  • Stephen J. McKenna
    • 2
  • Tracy A. Valentine
    • 1
  1. 1.Scottish Crop Research InstituteDundeeUK
  2. 2.School of ComputingUniversity of DundeeDundeeUK
  3. 3.Division of Civil EngineeringUniversity of DundeeDundeeUK
  4. 4.Earth and Life InstituteUniversité Catholique de LouvainLouvain-la-NeuveBelgium

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