Rice ratooning produces a ratoon crop (RC) with regenerated tillers in different nodes on the stubble of main crop (MC). It is crucial to apply nitrogen (N) at 15 d after heading and 1–2 d after harvest of MC for promoting bud initiation (N B ) and tiller growth (N T ), respectively to achieve high grain yield of RC. However, limited information is available on the N response of regenerated tillers in different node positions of RC. The hypothesis is that N response varied among node positions of regenerated tillers. [METHODS] Field experiments were conducted in 2019 and 2020 to compare yield performance of regenerated tillers of upper and lower node positions in RC under various combinations of N B and N T treatments and two cutting heights (15 and 45 cm) using a rice variety Huanghuazhan. showed that there was no significant difference in grain yield among N treatments of regenerated tillers in upper node (3rd and 2nd nodes from the top for 15 and 45 cm cutting height, respectively). However, N B and N T significantly increased the grain yield of regenerated tillers in lower nodes by 23.5 %− 52.7 % and 49.3 %− 115.7 %, respectively, which was mainly attributed to increased panicles m −2 for both cutting heights. N application also increased total dry weight and total N uptake of regenerated tillers in lower nodes, but not in upper node. Furthermore, a significant and positive correlation between grain yield and total N uptake was observed for regenerated tillers in lower nodes, but not in upper node. Our results indicate that regenerated tillers in lower nodes were more responsive to N application than those in upper node. [IMPLICATIONS] Therefore, optimization of N management for RC should emphasize on the N response of regenerated tillers in lower nodes for achieving high grain yield of RC. • Nitrogen (N) responses of tillers varied among node positions in ratoon crop. • Nitrogen responses of tillers in lower nodes were higher than in upper node. • Cutting height did not change N responses of tillers in various node positions. • Yield variation of ratoon crop was mainly due to the tillers in lower nodes.