A pot experiment using the root separation technique was carried out to study effect of root interaction in soybean maize intercropping with or without N fertilization. Results of the experiment show that the chlorophyll content of soybean was higher in Treatment NS (No separation) than in Treatment PS (partially separated with nylon net) and Treatment FS (fully separated) at all the growth stages with or without N fertilization. N fertilization was better at improving agronomic traits of the soybean, such as plant height, node number of the main stem, effective branches, pods per plabt and 100-seed weight, than mere intercropping (without N fertilization), and root interaction between the two interplanted crops played a significant（p＜0.05） role only in improving pods per plant and 100-seed weight of the soybean. The treatments of root separation underground also demonstrated that the root interaction improved economic and biological yields of the soybean, but the effect was not significant than that of N fertilization. Intercropping and N fertilization could not only increase the populations of bacteria, fungi, actinincrease and nitrogen-fixing bacteria, but also enhanced the activities of urease, phosphatase, invertase and protease. Correlation analysis revealed that the populations of bacteria, fungi, actinincrease, and nitrogen-fixing bacteria and the activities of urease, phosphatase, invertase and protease, were all significant (p＜0.05) and markedly significant (p＜0.01) related to chlorophyll content of soybean and economic and biological yields of soybean per plant. All the findings demonstrate that root interaction between interplanted crops plays an important role in improving soil micro-ecoenvironment, increasing the population of microorganisms and the activity of enzymes in the soil, and raising economic and biological yields of the crops.