A New Cereal Seed Treatment Method for Displaying Endosperm Cell Structures Under Micro CT Scanning

doi:10.11983/CBB24022

Abstract

Abstract: Cereal starch endosperm is the main source of human staple food, but the methods for observing its mature cell structure are not well developed. Micro CT technology, a non-destructive three-dimensional imaging technique, is a powerful tool for studying plant morphology. However, due to the uniform density of cereal starch endosperm, whose cell structures cannot be distinguished clearly by conventional micro CT techniques. In this study, we used phosphotungstic acid to treat ten different kinds of cereal seeds of seven crops, and then dried them by CO₂ critical point dryer. We found that the cell structure of the treated endosperms can be clearly displayed by micro CT. Our method provides a new way for studying the structures and functions of cereal starch endosperm cells.

Key words: micro CT, cereal starch endosperm, phosphotungstic acid

Xiuping Xu, Xiaoyu Yang, Min Feng. A New Cereal Seed Treatment Method for Displaying Endosperm Cell Structures Under Micro CT Scanning[J]. Chinese Bulletin of Botany, 2025, 60(1): 81-89.

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URL: https://www.chinbullbotany.com/EN/10.11983/CBB24022

https://www.chinbullbotany.com/EN/Y2025/V60/I1/81

Figures/Tables 12

Table 1 Scanning voltage and resolution of ripe seeds

Seed	Voltage (kV)	Resolution (µm)
Rice	100	0.88
Rice (partially enlarged)	50	0.56
Purple rice	100	0.95
Red rice	100	0.74
Glutinous rice	100	0.88
Millet	41	0.68
Oats	100	0.88
Sorghum	100	1.15
Job’s tears	100	2.60
Corn	100	1.48
Corn (partially enlarged)	49	0.56
Wheat	100	1.15

Figure 1 Micro CT scanning of rice seeds by different treatments (A) Rice seed treated by FAA; (B) Rice seed treated by 10% cesium iodide; (C) Rice seed treated by 10% phosphotungstic acid; (D) Enlarged view of rice seed treated by 10% phosphotungstic acid. Bars=500 µm

Figure 2 Micro CT scanning of purple rice seeds by different treatments (A) Purple rice seed treated by FAA; (B) Purple rice seed treated by 10% cesium iodide; (C) Purple rice seed treated by Lugols solution; (D) Purple rice seed treated by 10% phosphotungstic acid; (E) Enlarged view of purple rice seed treated by 10% phosphotungstic acid. Bars=400 µm

Figure 3 Micro CT scanning of red rice seeds by different treatments (A) Red rice seed treated by FAA; (B) Red rice seed treated by 10% phosphotungstic acid. Bars=400 µm

Figure 4 Micro CT scanning of polished glutinous rice seeds by different treatments (A) Polished glutinous rice seed treated by FAA; (B) Polished glutinous rice seed treated by 10% phosphotungstic acid. Bars=400 µm

Figure 5 Micro CT scanning of millet seeds by different treatments (A) Millet seed treated by FAA; (B) Millet seed treated by 10% phosphotungstic acid. Bars=300 µm

Figure 6 Micro CT scanning of oat seeds by different treatments (A) Oat seed treated by FAA; (B) Oat seed treated by 10% phosphotungstic acid. Bars=500 µm

Figure 7 Micro CT scanning of sorghum seeds by different treatments (A) Sorghum seed treated by FAA; (B) Sorghum seed treated by 10% phosphotungstic acid. Bars=500 µm

Figure 8 Micro CT scanning of the seeds of Job’s tears by different treatments (A) The seed of Job’s tears treated by FAA; (B) The seed of Job’s tears treated by 10% phosphotungstic acid. Bars=500 µm

Figure 9 Micro CT scanning of maize seeds by different treatments (A) Maize seed treated by FAA; (B) Maize seed treated by 10% phosphotungstic acid; (C) Part of maize seed treated by 10% phosphotungstic acid; (D) Enlarged view of part of maize seed treated by 10% phosphotungstic acid. Bars=400 µm

Figure 10 Micro CT scanning of wheat seeds by different treatments (A) Wheat seed treated by FAA; (B) Wheat seed treated by 10% phosphotungstic acid. Bars=500 µm

Table 2 Effect of cereal seed endosperm by phosphotungstic acid treatment

Kind of seeds	Rice	Purple rice	Red rice	Glutinous rice	Millet (remove seed coat)	Oats	Sorghum (remove seed coat)	Job’s tears	Corn (remove seed coat)	Wheat
Treatment time (day)	40	40	40	40	30	60	70	90	90	180
Valid or not	Valid	Valid	Valid	Valid	Valid	Valid	Valid	Valid	Valid	Invalid

References 15

[1]	Becraft PW, Gutierrez-Marcos J (2012). Endosperm development: dynamic processes and cellular innovations underlying sibling altruism. WIREs Dev Biol 1, 579-593. DOI PMID
[2]	Hu J, Guan YJ (2022). Seed Biology, 2nd edn. Beijing: Higher Education Press. pp. 62-63. (in Chinese)
	胡晋, 关亚静 (2022). 种子生物学(第2版). 北京: 高等教育出版社. pp. 62-63.
[3]	Huang LJ, Fu WL (2021). A water drop-shaped slingshot in plants: geometry and mechanics in the explosive seed dispersal of Orixa japonica (Rutaceae). Ann Bot 127, 765-774.
[4]	Kaestner A, Schneebeli M, Graf F (2006). Visualizing three-dimensional root networks using computed tomography. Geoderma 136, 459-469.
[5]	Knipfer T, Reyes C, Earles JM, Berry ZC, Johnson DM, Brodersen CR, McElrone AJ (2019). Spatiotemporal coupling of vessel cavitation and discharge of stored xylem water in a tree sapling. Plant Physiol 179, 1658-1668. DOI PMID
[6]	Korte N, Porembski S (2012). A morpho-anatomical characterisation of Myrothamnus moschatus (Myrothamnaceae) under the aspect of desiccation tolerance. Plant Biol 14, 537-541.
[7]	Liao H, Fu XH, Zhao HQ, Cheng J, Zhang R, Yao X, Duan XS, Shan HY, Kong HZ (2020). The morphology, molecular development and ecological function of pseudonectaries on Nigella damascena (Ranunculaceae) petals. Nat Commun 11, 1777.
[8]	Lin JX, Hu ZJ, Zhang X, Yang SY, Shan XY (2019). Method for Improving Imaging Quality of Arabidopsis thaliana Seeds. Chinese Patent, CN109142398A. 2019-01-04. (in Chinese)
	林金星, 胡子建, 张曦, 杨舜垚, 单晓昳 (2019). 一种提高拟南芥种子成像质量的方法. 中国专利, CN109142398A. 2019-01-04.
[9]	Losso A, Bär A, Dämon B, Dullin C, Ganthaler A, Petruzzellis F, Savi T, Tromba G, Nardini A, Mayr S, Beikircher B (2019). Insights from in vivo micro-CT analysis: testing the hydraulic vulnerability segmentation in Acer pseudoplatanus and Fagus sylvatica seedlings. New Phytol 221, 1831-1842.
[10]	Perret JS, Al-Belushi ME, Deadman M (2007). Non-destructive visualization and quantification of roots using computed tomography. Soil Biol Biochem 39, 391-399.
[11]	Snell P, Wilkinson M, Taylor GJ, Hall S, Sharma S, Sirijovski N, Hansson M, Shewry PR, Hofvander P, Grimberg Å (2022). Characterisation of grains and flour fractions from field grown transgenic oil-accumulating wheat expressing oat WRI1. Plants 11, 889.
[12]	Staedler YM, Masson D, Schönenberger J (2013). Plant tissues in 3D via X-ray tomography: simple contrasting methods allow high resolution imaging. PLoS One 8, e75295.
[13]	Xu XP, Meng SC, Liang RH, Jin WQ, Feng M (2021). New method of improving micro-CT images contrasts on plant samples. J Chin Electron Microsc Soc 40, 460-466. (in Chinese)
	徐秀苹, 孟淑春, 梁荣花, 靳婉青, 冯旻 (2021). 磷钨酸和干燥处理提高植物样品显微CT成像对比度的方法. 电子显微学报 40, 460-466.
[14]	Zhang CQ, Li Y (2019). Crop Seed Science, 2nd edn. Beijing: China Agriculture Press. pp. 16-18. (in Chinese)
	张春庆, 李岩 (2019). 作物种子学(第2版). 北京: 中国农业出版社. pp. 16-18.
[15]	Zhang X, Hu ZJ, Guo YY, Shan XY, Li XJ, Lin JX (2020). High-efficiency procedure to characterize, segment, and quantify complex multicellularity in raw micrographs in plants. Plant Methods 16, 100. DOI PMID