Abstract: Soybean plants are able to develop resistance against Heterodera glycines when coated with Bacillus simplex. Three bio-control bacteria were selected after a two-year field experiment. Among them, strain Sneb545 was proven effective by pot culture in greenhouse. In order to elucidate the modes of action, the split-root experiment was designed to investigate Sneb545's ability to induce soybean systemic resistance against H. glycines. The results showed that soybean seeds treated with strain Sneb545 were able to produce seedlings that had notable resistance to first generation cyst nematodes. Number of H. glycines cyst in soybean roots decreased by 72.63% comparing with control, the ratio of H. glycines in adjacent soil to soybean root decreased by 70.63%. Split-root experiment concluded that after inoculating Sneb545 in the challenging roots, nematodes penetration in responding roots decreased by 51.27%, number of cyst in soil decreased by 65.82%. Sneb545 was identified as B. simplex based on morphological, physiological, biochemical characteristics and 16S rDNA sequence analysis.