ChronoRoot 2.0: An Open AI-Powered Platform for 2D Temporal Plant Phenotyping

The analysis of plant developmental plasticity, including root system architecture, is fundamental to understanding plant adaptability and development, particularly in the context of climate change and agricultural sustainability. While significant advances have been made in plant phenotyping technologies, comprehensive temporal analysis of root development remains challenging, with most existing solutions providing either limited throughput or restricted structural analysis capabilities. Here, we present ChronoRoot 2.0, an integrated open-source platform that combines affordable hardware with advanced artificial intelligence to enable sophisticated temporal plant phenotyping. The system introduces several major advances, offering an integral perspective of seedling development: (i) simultaneous multi-organ tracking of six distinct plant structures, (ii) quality control through real-time validation, (iii) comprehensive architectural measurements including novel gravitropic response parameters, and (iv) dual specialized user interfaces for both architectural analysis and high-throughput screening. We demonstrate the system's capabilities through three use cases for Arabidopsis thaliana: characterization of circadian growth patterns under different light conditions, detailed analysis of gravitropic responses in transgenic plants, and high-throughput screening of etiolation responses across multiple genotypes. ChronoRoot 2.0 maintains its predecessor's advantages of low cost and modularity while significantly expanding its capabilities, making sophisticated temporal phenotyping more accessible to the broader plant science community. The system's open-source nature, combined with extensive documentation and containerized deployment options, ensures reproducibility and enables community-driven development of new analytical capabilities.