A high throughput method for quantifying number and size distribution of Arabidopsis seeds using large particle flow cytometry

Plant Methods
Alejandro MoralesM van Zanten

Abstract

Seed size and number are important plant traits from an ecological and horticultural/agronomic perspective. However, in small-seeded species such as Arabidopsis thaliana, research on seed size and number is limited by the absence of suitable high throughput phenotyping methods. We report on the development of a high throughput method for counting seeds and measuring individual seed sizes. The method uses a large-particle flow cytometer to count individual seeds and sort them according to size, allowing an average of 12,000 seeds/hour to be processed. To achieve this high throughput, post harvested seeds are first separated from remaining plant material (dust and chaff) using a rapid sedimentation-based method. Then, classification algorithms are used to refine the separation process in silico. Accurate identification of all seeds in the samples was achieved, with relative errors below 2%. The tests performed reveal that there is no single classification algorithm that performs best for all samples, so the recommended strategy is to train and use multiple algorithms and use the median predictions of seed size and number across all algorithms. To facilitate the use of this method, an R package (SeedSorter) that implements the met...Continue Reading

References

Feb 24, 2009·Trends in Plant Science·Melissa A FitzgeraldRobert D Hall
Apr 22, 2010·The New Phytologist·Ada LinkiesGerhard Leubner-Metzger
Nov 1, 1992·Trends in Ecology & Evolution·M WestobyM Leishman
Feb 10, 2011·Plant Methods·Rowan P HerridgeRichard C Macknight
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Oct 23, 2012·Plant Physiology·Thu-Phuong NguyenLeónie Bentsink
Jun 19, 2013·Plant Physiology·Wen-Bo JiangWen-Hui Lin
Jul 18, 2015·Journal of Experimental Botany·Thu-Phuong NguyenLeónie Bentsink
Sep 25, 2016·Plant Physiology·Siegfried JahnkeHanno Scharr
Apr 6, 2018·Genetics·Andrew W Woodward, Bonnie Bartel

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Methods Mentioned

BETA
flow cytometry
transgenic
Fluorescence

Software Mentioned

R
InterGenotype
phenoSeeder
BioSorter
mlr
FlowPilot
IntraPlant
R package
SeedSorter
SeedSorter R

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