Soybean cyst nematode (SCN, ) is an obligate sedentary biotroph that poses major threats to soybean production globally. Recently, multiple miRNAome studies revealed that miRNAs participate in complicated soybean-SCN interactions by regulating their target genes. However, the functional roles of miRNA and target genes regulatory network are still poorly understood. In present study, we firstly investigated the expression patterns of miR159 and targeted genes. The results showed downregulation during SCN infection; conversely, genes upregulated. Furthermore, miR159 overexpressing and silencing soybean hairy roots exhibited strong resistance and susceptibility to , respectively. In particular, miR159- genes are reported to be involve in GA signaling and metabolism. Therefore, we then investigated the effects of GA application on the expression of module and the development of . We found that GA directly controls the miR159- module, and exogenous GA application enhanced endogenous biologically active GA and GA, the abundance of miR159, lowered the expression of genes and delayed the development of . Moreover, SCN infection also results in endogenous GA content decreased in soybean roots. In summary, the soybean miR159- module was directly involved in the GA-modulated soybean resistance to .