PAT (Periderm Assessment Toolkit): A Quantitative and Large-Scale Screening Method for Periderm Measurements.

Abstract

The periderm is a vital protective tissue found in the roots, stems, and woody elements of diverse plant species. It plays an important function in these plants by assuming the role of the epidermis as the outermost layer. Despite its critical role for protecting plants from environmental stresses and pathogens, research on root periderm development has been limited due to its late formation during root development, its presence only in mature root regions, and its impermeability. One of the most straightforward measurements for comparing periderm formation between different genotypes and treatments is periderm (phellem) length. We have developed PAT (Periderm Assessment Toolkit), a high-throughput user-friendly pipeline that integrates an efficient staining protocol, automated imaging, and a deep-learning-based image analysis approach to accurately detect and measure periderm length in the roots of Arabidopsis thaliana . The reliability and reproducibility of our method was evaluated using a diverse set of 20 Arabidopsis natural accessions. Our automated measurements exhibited a strong correlation with human-expert-generated measurements, achieving a 94% efficiency in periderm length quantification. This robust PAT pipeline streamlines large-scale periderm measurements, thereby being able to facilitate comprehensive genetic studies and screens. Although PAT proves highly effective with automated digital microscopes in Arabidopsis roots, its application may pose challenges with nonautomated microscopy. Although the workflow and principles could be adapted for other plant species, additional optimization would be necessary. While we show that periderm length can be used to distinguish a mutant impaired in periderm development from wild type, we also find it is a plastic trait. Therefore, care must be taken to include sufficient repeats and controls, to minimize variation, and to ensure comparability of periderm length measurements between different genotypes and growth conditions.

Plant phenotyping relevance

植物根の表現型（周皮長）を自動画像取得・深層学習解析で定量するパイプラインを開発し、専門家測定との相関で信頼性と再現性を検証しており、方法が研究の中心である。

Code and data availability

The authors publicly release the PAT pipeline (analysis code/scripts) and a test dataset of Col-0 and wox4-1 TIFF microscopy images via their GitHub repository. Full-resolution TIFF images of the 20 natural accessions are only available upon request (request_only, not listed as an allowed URL).