To explore the interaction effect of different nitrogen levels and mepiquat chloricie （DPC） on cotton， a two-factor cleft zone test was designed with “XinLuzhong 88” as test material， two nitrogen levels of 320 （N₁）， 480 kg/hm² （N₂）， and four doses of DPC spray of 67.0 （F1）， 150.0 （F2）， 260.5 （F3）， 371.0 g/hm² （F4）， were applied， the effect of the DPC under different nitrogen levels on photosynthetic characteristics and yield formation was studied. The results showed that there were interactions between nitrogen level and DPC dosage on the net photosynthetic rate （P_n）， transpiration rate （T_r）， dry matter accumulation， nitrogen accumulation and yield composition factors in cotton. The promotion effects of nitrogen fertilizer and DPC on P_n and T_r of cotton were mainly in cotton after full boll and opening boll stages， and both P_n and T_r were higher under N₁F3 treatment. The accumulation of dry matter， nitrogen accumulation and lint yield of final reproductive organs in N₁ treatment were higher than that of N₂， and the indexes were 5.6%， 11.32% and 7.68% higher respectively. Under the same nitrogen level， the accumulation of dry matter and nitrogen in reproductive organs， the number of single knots， single bell mass and lint yield of cotton， were significantly increased with N₁F3 treatment， but it did not reach a significant difference level compared with N₁F2 treatment， among which the lint yield of N₁F3 was 2 803.32 kg/hm²， only 0.3% higher than that of N₁F2 treatment. Therefore， considering the performance of each measurement index， under the test conditions， it is recommended to apply a nitrogen amount of 320 kg/hm²， and a DPC dosage of 150.0 g/hm² to ensure a higher cotton yield.