Unreasonable soil tillage measures have caused a sharp decline in the soil carbon (C) pool capacity of rice (Oryza sativa L.) paddy fields, have reduced soil fertility, and have threatened the safe production of rice. Based on long-term position–location experiments started in 2005, this paper systematically studied the effects of different soil tillage treatments (CT: no return of plowing straw to the field as control; CTS: return of plowing straw to the field; NTS: return of no-tillage straw to the field; RTS: return of rotary plowing straw to the field) on soil physical and chemical properties and soil organic carbon (SOC) accumulation characteristics in rice paddy fields, in order to clarify the impact of different long-term soil tillage measures on soil carbon cycle microecology in double-cropped rice paddy fields and provide a theoretical basis for soil SOC sequestration and the sustainable utilization of rice paddy fields in double-cropped rice paddy fields in southern China. The results were as follows: A total of 30.7–40.7% of the SOC stored in rice paddy fields was residue C derived from microorganisms, and 45.7–54.2% of SOC accumulation came from plant residue-derived C. Straw return treatments (CTS, RTS, and NTS) significantly increased soil lignin phenol content and promoted the accumulation of plant-derived SOC. Soil lignin phenol content in the RTS treatment was significantly higher than that in the CTS treatment (p < 0.05). Amino sugar content in rhizosphere soil was higher than that in non-rhizosphere soil. The measure of returning straw to the field increased amino sugar content in the rhizosphere and non-rhizosphere. C derived from plants was greater than that from microbial residues in double-cropped rice paddy fields in southern China. Hence, no-till/rotary tillage and straw return can improve the sequestration of soil SOC, which is of great significance for achieving “C neutrality” and alleviating the pressure on food security.