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Summary  Root analysis is essential for both academic and agricultural research. Despite the great advances in root phenotyping and imaging, calculating root length is still performed manually and involves considerable amounts of labor and time. To overcome these limitations, we developed My ROOT  ,  a software for the semiautomatic quantification of root growth of seedlings growing directly on agar plates. Our method automatically determines the scale from the image of the plate, and subsequently measures the root length of the individual plants. To this aim, My ROOT  combines a bottom‐up root tracking approach with a hypocotyl detection algorithm. At the same time as providing accurate root measurements, My ROOT  also significantly minimizes the user intervention required during the process. Using Arabidopsis, we tested My ROOT  with seedlings from different growth stages and experimental conditions. When comparing the data obtained from this software with that of manual root measurements, we found a high correlation between both methods ( R  2  = 0.997). When compared with previous developed software with similar features ( BRAT  and  EZ ‐Rhizo), My ROOT  offered an improved accuracy for root length measurements. Therefore, My ROOT  will be of great use to the plant science community by permitting high‐throughput root length measurements while saving both labor and time.

My ROOT : a method and software for the semiautomatic measurement of primary root length in Arabidopsis seedlings