免疫金标记分析水稻细胞壁中的细胞壁多糖尤其是(1;3,1;4)-β-D-葡聚糖

Ryusuke Yokoyama; Natsumi Kido; Tsuyoshi Yamamoto; Jun Furukawa; Hiroaki Iwai; Shinobu Satoh; Kazuhiko Nishitani

doi:10.21769/BioProtoc.1748

Improve Research Reproducibility A Bio-protocol resource

提交稿件
订阅
CN
- EN - English
- CN - 中文

Peer-reviewed

Immunogold Labeling Analysis of Cell Wall Polysaccharides with Special Reference to (1;3,1;4)-β-D-glucan in Rice Cell Walls

免疫金标记分析水稻细胞壁中的细胞壁多糖尤其是(1;3,1;4)-β-D-葡聚糖

KN Kazuhiko Nishitani email

发布: 2016年03月05日第6卷第5期 DOI: 10.21769/BioProtoc.1748 浏览次数: 8976

评审: Tie LiuCarsten AdeValérie Cornuault

下载 PDF

Q&A

引用

Cited by

参见作者原研究论文

The authors used this protocol in:

Cover of Plant & Cell Physiology, featuring study using the protocol.

Feb 2015

实验方案合集

Cell Imaging - A Special Collection for Cell Bio 2023

相关实验方案

一种简便、快速且经济的方法评估微藻和拟南芥的碳水化合物分配

Araceli N. Bader [...] Leonardo Curatti

2024年12月05日 959 阅读

甜樱桃发育中花原基的淀粉酶法定量检测

Nestor Santolaria [...] Afif Hedhly

2025年04月05日 751 阅读

检测和分离拟南芥种子黏液中鼠李半乳糖醛酸Ⅱ的新方法

Dayan Sanhueza and Susana Saez-Aguayo

2025年09月05日 174 阅读

Abstract

Various types of cell wall compositions have evolved to fulfill a wide range of biological roles during the diversification of land plants. (1;3,1;4)-β-D-glucan (MLG) is a defining feature of the cell walls in the order Poales (Yokoyama and Nishitani, 2004), which has multiple functions associated with metabolic, growth, and defense systems. MLG is also a characteristic component of the matrix polysaccharides that undergo turnover and metabolism, depending on the tissue and the stage of development (Kido et al., 2015). Determining the extracellular localization of MLG is essential for elucidating its functions. Electron microscopy immunogold labeling analysis is a useful technique, which provides an accurate representation of the extracellular distribution of MLG. This strategy is also applicable to various kinds of cell wall polysaccharides, which have key roles in regulating growth and differentiation in each plant species.

Keywords: Electron microscopy (电子显微镜)

Cell wall (细胞壁)

Immunogold labeling (金标记)

Rice (大米)

Mixed-linkage glucan (混联葡聚糖)

Materials and Reagents

Gelatin capsule (HF capsule) (Matsuya Co.)
Rice plants (see Note)
Paraformaldehyde (Electron Microscopy Sciences, catalog number: 12300 )
Sodium cacodylate buffer solution (TAAB Laboratories BATCH, catalog number: 90760 )
Glutaraldehyde (Electron Microscopy Sciences, catalog number: 16220 )
Tannic acid (Sigma-Aldrich, catalog number: 403040 )
Phosphate buffer powder (Siyaku, Wako Pure Chemical Industries, catalog number: 167-14491 )
Ethanol (Siyaku, Wako Pure Chemical Industries, catalog number: 057-00456 )
LR White resin (Electron Microscopy Sciences, London Resin Company, catalog number: 14381-CA )
Monoclonal antibody against (1;3,1;4)-β-D-glucan (mouse IgG, Kappa Light) (Biosupplies, catalog number: 400-3 )
Note: It is named as (1-3;1-4)-β-D-glucan on Biosupplies’ website.
Goat Anti-mouse IgG pAb, Gold 15 nm, EM (BBI Solutions, catalog number: EM.GMHL15 )
Phosphate-buffered salts (Takara Bio Company, catalog number: T900 )
Bovine serum albumin (BSA) (Sigma-Aldrich, catalog number: A2058 )
Uranyl acetate (Siyaku, Wako Pure Chemical Industries, catalog number: 554-85072 )
Lead stain solution (Sigma-Aldrich, catalog number: 18-0875-2-25 ml-J )
Note: This reagent may not be available outside Japan, and can be replaced with alternative lead stains. For an alternative, readers are referred to a more general experimental manual for electron microscopy of plant cells (Hall and Hawes, 1991).
Molecular sieves (Sigma-Aldrich, catalog number: M6141 )
Fixative solution (see Recipes)
2% glutaraldehyde solution (see Recipes)
100% ethanol solution (see Recipes)
1x Phosphate-buffered saline (PBS) (see Recipes)

Equipment

Growth chamber (Nippon Medical & Chemical Instruments, model: LH220S )
Ultracut UCT Ultramicrotome (Leica Microsystems)
Diaphragm vacuum pump (Leybold-Heraus, model: Divac 2.2 L )
Desiccator (Sanplatec, model: PC-250K )
Grid mesh (Okenshoji, catalog number: 09-1079 )
Transmission electron microscope (JEOL USA, model: JEM-1400plus )
CCD camera (Olympus soft imaging solutions GmbH, model: Veleta )

Procedure

English

中文翻译

文章信息

版权信息

如何引用

Yokoyama, R., Kido, N., Yamamoto, T., Furukawa, J., Iwai, H., Satoh, S. and Nishitani, K. (2016). Immunogold Labeling Analysis of Cell Wall Polysaccharides with Special Reference to (1;3,1;4)-β-D-glucan in Rice Cell Walls. Bio-protocol 6(5): e1748. DOI: 10.21769/BioProtoc.1748.