Abstract: 【Objective】The purpose of this study was to explore the response of soil nitrogen cycling microbes and the functional genes to biochar,and to clarify the mutual feedback mechanism between soil physicochemical properties and nitrogen cycling microbes,so as to provide theoretical basis for improving soil,improving crop nitrogen use efficiency and regulating soil nitrogen cycle process.【Method】The effects of biochar on nitrogen accumulation in plants, the physicochemical properties of tobacco rhizosphere soil,the relative abundance of nitrogen cycling microor-ganisms,the expression of their functional gene were analyzed by a combination of metagenomics and qRT-PCR using the variety Yunyan 87as the test material,comparing the biochar improvement by soil tillage layer(T1,30 t/ha)and no biochar application(T0,control).【Result】Compared to control T0,nitrogen accumulation in flue-cured tobacco was significantly increased by45.75% and 29.91% at T1 treatment in 2021 and 2022,respectively(P<0.05,the same below). At the same time, soil alkali-hydrolyzed nitrogen and nitrate nitrogen content in T1 treatment were significantly increased, while ammonium nitrogen content was significantly decreased compared to control T0 treatment. Metagenomics analysis showed that T1 treatment significantly increased the relative abundances of Thaumarchaeota,Acidobacteriota,Chloroflexi and Nitrospirae at the phylum level in rhizosphere soil compared to T0 treatment,among which the relative abundance of Thaumarchaeota increased by 23.76%. At the genus level,the relative abundance of Candidatus_Nitrosotalea in T1 treatment was significantly increased by 20.83% compared to T0 treatment. In terms of nitrogen cycle function subcategories,T1 treatment significantly increased the relative expression of nitrification nxrA,ammonia oxidation amoA and nitrogen fixation nifA genes in the process of soil nitrogen cycle by 20.08%,250.37% and 82.45% respectively,compared to T0 treatment. While reducing the relative expression of denitrification norB genes by 39.76%. qRT-PCR analysis results showed that the addition of biochar had a certain persistence in the change of soil nitrogen cycle process. The further redundancy analysis found that soil pH,inorganic nitrogen content,bulk density,porosity and other physical properties were important environmental factors affecting soil nitrogen cycle. Pearson correlation analysis found that soil pH,nitrate nitrogen and physical properties were significantly positively correlated with nitrification and ammonia oxidizing microbes abundance and genes expression(P<0.01).【Conclusion】The improvement of tobacco planting soil with 30 t/ha biochar is beneficial to improve soil physicochemical properties and then drive the transformation of soil nitrogen cycle to efficient nitrogen utilization pathway,thereby promoting nitrogen absorption by plants in tobacco-rice cropping area.