Investigating the biochar and functional fungal agents to coordinate the regulation of desertified soil nutrients and microbial diversity and function can provide a theoretical basis for the fertility cultivation of desertified soil. The soil of the typical ecological and economic forest of the Mu Wusu Sandy Land was selected as the research object, and the effects of different amounts (2%, 4%, 8%) of biochar treatment and its combined application of quantitative functional agent treatment (Bacillus subtilis + Bacillus megaterium + Bacillus colloid) on soil chemical properties, bacterial diversity and function were analyzed by field positioning experiment and high-throughput amplification and sequencing technology. The results showed that: (1) Compared with the single biochar treatment, the soil available nitrogen and phosphorus nutrient contents of the biochar + functional agent treatment group increased by 44.71% and 187.36%, respectively. (2) The relative abundance of the Acidobacteria was significantly reduced by the single application or combination of functional agents of biochar, while the relative abundance of the phylum Firmcutes in the treatment of biochar + functional bacteria increased by 163.80%, 155.15%, and 100.21%, respectively, and the richness and diversity of soil bacterial species were increased to varying degrees. (3) The combined application of functional agents and biochar changed the carbon and nitrogen cycle function mediated by soil bacteria, such as significantly improving the decomposition function of organic matter of soil bacteria, and the decomposition process of chitosan and xylan increased by 186.54% and 242.46% respectively compared with the control group; enhanced the ammonization and respiratory function of bacteria; while single biochar treatment improved the denitrification and nitrogen fixation function of the bacteria. In summary, biochar with functional bacterial agents is better than single application of biochar treatment. Biochar and functional bacteria agents added to improve the soil available nitrogen and phosphorus nutrient content, significantly changed the soil bacterial community diversity and community function. These results can provide a scientific basis and theoretical reference for the selection of desertification soil improvement methods.