Soil organic carbon is essential for maintaining terrestrial ecosystem function and mitigating soil degradation. Soil microorganisms participate in soil carbon cycling. They are affected by tillage methods and straw returning. A split-plot design was adopted in this experiment. The whole-plot treatment had two tillage methods, subsoil tillage (ST) and rotary tillage (RT). The split-plot treatment included full straw returning (F) and no straw returning (0). The microbial community structure and carbon sequestration genes were assessed by Illumina sequencing technique. Soil organic carbon contents were measured during 2012-2017. The results showed that 1) subsoil tillage and straw returning significantly increased pH, microbial biomass carbon, total nitrogen, silt content, and clay content, while significantly decreased sand content; 2) during the test period (2012-2017), soil organic carbon (SOC) content under all treatments showed an increasing trend, but the increment for average SOC content under straw returning and subsoiling treatments was significantly higher than that of no straw returning and rotary tillage by 33.2 % and 30.6%, respectively; 3) Proteobacteria was the most abundant type of bacteria in the soil, followed by Acidobacteria and Gemmatadanetes; 4) STF treatment maintained high microbial diversity; 5) Excepted for soil sand content, soil pH, microbial biomass carbon, total nitrogen, silt content and clay content all caused the variation of soil microbial community structure under the STF treatment in the direction of SOC accumulation; 6) in addition to the gene abundance in the di- and oligosaccharides metabolic pathway, the gene abundance in the metabolic pathways for CO fixation, central carbohydrate metabolism, fermentation, one-carbon metabolism, organic acids, sugar alcohols and glycoside hydrolases showed that subsoil tillage was significantly higher than rotary tillage, with posi-tively correlation with soil organic carbon content. Therefore, the combination of subsoil tillage and straw returning could improve basic soil properties, affect soil microbial community structure, and increase the capacity of soil carbon fixation, thus providing a realistic basis for solving soil degradation.