By investigating the factors influencing the transformation efficiency of Agrobacterium, a genetic transformation system of soybean cotyledonary node with non tissue-culture was optimized according to the transient expression of GUS in the region of cotyledonary node; The gene Bar was transformed into soybean jidou16, and the transgenic plants were screened by glufosinate injected on leaves. The results indicated that infection medium contains 3% sugar and none Silwet L-77、OD6OO=0.6、with the absorbent cotton as attached media、infect one time have the highest percentage of GUS positive, was 62.13%. We obtained 10 positive plants in T0 generation after detected the transgenic plants resistant to glufosinate by PCR, the transformation efficiency was 2.5%. 3 positive plants of T1 generation were obtained by detection of PCR and RT-PCR, preliminary demonstrated that the target gene was integrated into the soybean genome.