Masticatory hypofunction and soft food affect the tooth rows, occlusion, and jawbone. This study aimed to clarify the influence of tooth loss and a soft diet <span>on morphology of the tooth root during the growth period. We divided</span><span> 3-week-</span><span>old Wistar rats into the following three groups: Hard diet group (rats raised on solid standard diet), Powder diet group (rats raised on powdered standard feed diet), and Extraction group (rats raised on powdered standard diet with maxillary molars extraction). Length, width, cross-sectional area, and volume </span><span>of the root of the mandibular M1 and M2 were measured using micro-CT</span><span> analysis. Non-decalcified thin-slice specimens of sagittal sections of the M1 were obtained at the age of 20 weeks, and the roots were observed. The root length of all roots in the Extraction group was significantly longer than that in the other groups. The root width and cross-sectional area at the apical side 1/4 of all roots in the Extraction group were significantly smaller than those in the other groups. The root volume of the M1 mesial root in the Extraction group was significantly smaller than that in the other groups.</span><span> </span><span>This study clarified that when masticatory stimulus in the immature teeth is reduced by the extraction of opposing teeth and a powder diet, the root length increases due </span><span>to the promotion of cellular cementum addition at the apex, and the root</span><span> width and cross-sectional area decrease due to the suppression of cellular cementum addition at the apical side 1/4 of the roots.</span>