Gypsum is already a well-known soil conditioner, especially in regions with predominantly acidic soils. More recently gypsum has also been recognized as a strategy to increase carbon (C) accumulation in the soil, thus contributing to reduced C emissions agriculture. However, there are still doubts regarding how gypsum influences soil C accumulation and the role of the root system in this process. The objective of the present study was to evaluate the influence of gypsum on C accumulation and the relationships of C with soil chemical properties and development of the sugarcane root system to a depth of 2 m, evaluated in the field with an Oxisol. The experiment was arranged in a randomized complete block design, with four replications and two treatments: control (without gypsum) and gypsum application (5 t ha(-1)). Sugarcane stalk biomass and straw production were evaluated in the first planted sugarcane and six ratoon crops. At 87 months after gypsum application, after the seventh cut, the soil was sampled in the depth of 0-200 cm, in 20 cm intervals, for evaluation of the root dry weight (RW) and soil chemical properties (S-SO42-), calcium (Ca2+), aluminum saturation (Al3+ saturation), pH, free fulvic acid carbon (FFAC), total C (TC) and particulate C (PC). Stocks of PC and TC were also estimated. The application of 5 t ha(-1) gypsum favored the increase of S-SO42- and Ca2+ contents and induced a reduction in Al3+ saturation in the soil profile. In association with these improvements in the chemical properties of the subsurface layers were increases in the RW, FFAC, TC and PC stocks. After 87 months of gypsum application, the TC and PC stocks increased by 12.3 and 2.2 t ha(-1), respectively, in the 0-200 cm layer, with the highest gains in the 40-200 cm layer. The multivariate analysis showed that in the 40-200 cm layer of this extremely weathered soil, the effect of gypsum on increasing TC and PC stocks, as well as FFAC, is a result of the reduced Al3+ saturation associated with the increase in nutrients such as Ca and S, stimulating greater root development.