Study on the effects of total straw incorporation combined with soil modified fertilizer and compound microbial agent on soil nutrition, soil physical structure, and soil microorganism through rice-wheat rotation in the field can provide practical basis and technical support for rapid soil fertilization and improve medium-and low-yield farmland production. An experiment was carried out in the Yellow River Bay Project Base of Binhai County, Yancheng City, Jiangsu Province. Five treatments were set up including ① modified organic substrate (MOS)+compound microbial agents (CMA)+chemical fertilizer (CF); ② CMA+CF; ③ MOS+CF; ④ CF; and ⑤ control check (CK). During the whole growth stage of wheat, soil chemical and physical properties, enzyme activities, microbial biomass carbon and nitrogen, and microbial diversity were analyzed with the soil samples origin from wheat greening (S1), wheat jointing and booting (S2), and wheat maturity (S3). The results showed that compared with CK, MOS+CMA+CF treatment could increase soil available nutrient content in a short time with available nitrogen, available phosphorus and available potassium increased by 23.59%, 40.74% and 43.78%, respectively, as well as decrease soil bulk density, increase soil porosity, and significantly increase soil microbial biomass nitrogen content and soil urease activity. MOS+CMA+CF could also increase the diversity and abundance of soil bacteria and fungi at the earlier wheat growth stage, and finally increase wheat yield (compared with CF and CK, wheat yield increased by 24.93% and 149.29%). CMA+CF could significantly improve soil cellulase activity and soil microbial biomass carbon content, indicating better straw degradation ability. MOS was rich in organic matter, which could maintain and improve soil organic matter content and improve soil physical environment. In general, MOS+CMA+CF treatment was the most significant treatment for improving soil fertility and increasing wheat yield in the short term. While CMA+CF treatment showed advantages in accelerating the degradation of returning straw and other soil properties. Compared with the large dosage and high price of MOS+CMA+CF treatment, CMA+CF treatment had a great application prospect to provide technical support for the development of medium-and low-yield farmland.