A Simple Protocol for Mapping the Plant Root System Architecture Traits

摘要

We use simple laboratory tools to examine the root system architecture (RSA) of Arabidopsis and Medicago. The plantlets are grown hydroponically over mesh and spread using an art brush to reveal the RSA. Images are taken using scanning or a high-resolution camera, then analyzed with ImageJ to map traits.

摘要

Comprehensive knowledge of plant root system architecture (RSA) development is critical for improving nutrient use efficiency and increasing crop cultivar tolerance to environmental challenges. An experimental protocol is presented for setting up the hydroponic system, plantlet growth, RSA spreading, and imaging. The approach used a magenta box-based hydroponic system containing polypropylene mesh supported by polycarbonate wedges. Experimental settings are exemplified by assessing the RSA of the plantlets under varying nutrient (phosphate [Pi]) supply. The system was established to examine the RSA of Arabidopsis, but it is readily adaptable to study other plants like Medicago sativa (Alfalfa). Arabidopsis thaliana (Col-0) plantlets are used in this investigation as an example to understand the plant RSA. Seeds are surface sterilized by treating ethanol and diluted commercial bleach, and kept at 4 °C for stratification. The seeds are germinated and grown on a liquid half-MS medium on a polypropylene mesh supported by polycarbonate wedges. The plantlets are grown under standard growth conditions for the desired number days, gently picked out from the mesh, and submersed in water-containing agar plates. Each root system of the plantlets is spread gently on the water-filled plate with the help of a round art brush. These Petri plates are photographed or scanned at high resolution to document the RSA traits. The root traits, such as primary root, lateral roots, and branching zone, are measured using the freely available ImageJ software. This study provides techniques for measuring plant root characteristics in controlled environmental settings. We discuss how to (1) grow the plantlets, and collect and spread root samples, (2) obtain pictures of spread RSA samples, (3) capture the images, and (4) use image analysis software to quantify root attributes. The advantage of the present method is the versatile, easy, and efficient measurement of the RSA traits.

引言

The root system architecture (RSA), which is underground, is a vital organ for plant growth and productivity^1,2,3. After the embryonic stage, plants undergo their most significant morphological changes. The way in which the roots grow in the soil greatly affects the growth of plant parts above ground. Root growth is the first step in germination. It is an informative trait as it uniquely responds to different available nutrients^1,2,3,4. The RSA exhi....

方案

The whole protocol is summarized schematically in Figure 1, showing all the essential steps involved in revealing the root system architecture (RSA) of plantlets. Protocol steps are given in detail below:

1. Arabidopsis seed surface sterilization

1. Transfer a tiny scoop (approximately 100 seeds = approximately 2.5 mg) of seeds to a microfuge tube, and soak for 30 min in distilled water at room temperature (RT). This entire procedure is carried out in the aseptic condition.
2. Briefly centrifuge the microfuge tube containing seeds at 500 x g for 5 s, using any tabletop ....

结果

The different morphometric traits of root system architecture (RSA) are measured using simple laboratory tools, and the steps are depicted schematically in Figure 1. The details of the hydroponic setup demonstrate the protocol's potential in measuring the RSA (Figure 1 and Figure 2).

Given the observed differences in gelling agents, we used a hydroponic growing system to conduct all the studies

讨论

This work demonstrated mapping RSA utilizing simple laboratory equipment. Using this method, phenotypic alterations are recorded at the refined level. The benefit of this strategy is that the shoot portion never comes in contact with the media, so the phenotype of the plantlets is original. This method involves setting up a hydroponic system to grow plantlets as described in the protocol. Then, each plantlet is taken out intact and placed on an agar-filled Petri plate. The root system is then allowed to spread manually u.......

材料

本文使用的材料清单

姓名	公司	目录编号	评论
拟南芥 （Col 0）	Lehle 种子	WT-02	哥伦比亚 （Col-0**，无标记）*
艺术画笔	亚马逊或任何其他供应商		水彩圆刷尺寸 14 （8 毫米）、16 （9.5 毫米）、18 （12 毫米） 和 20 （14.2 毫米）
带数码相机的自动显微镜	徕卡显微系统		LAS 版本 4.12.0，徕卡显微系统
成像软件	ImageJ		ImageJ V  1.8.0
品红色盒子 GA-7	Fisher Scientific	50-255-176
紫花苜蓿	Johnny's 种子
培养皿 （150 mm x 15 mm）	美国科学公司	8609-0215	150 mm x 15 mm PS 培养皿 （https://www.usascientific.com）
照相相机	Cannon 或 Nikon		距模式下的任何高百万像素（每英寸至少 12 百万像素）相机
植物琼脂	Sigma-Aldrich	A3301	沉香胶 适用于植物组织培养
聚碳酸酯片	Amazon		1毫米  厚
聚丙烯网	Amazon		孔径250 µm、500 和微型;m 和 1000 µm
扫描仪	Epson		Epson Perfection V700 照片 （扫描速度 600 dpi）