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The Antisense 4CL gene was transfected into alfalfa through Agrobacterium-mediated transfer. The test results indicated that the antisense 4CL gene was successfully integrated into the genome DNA of alfalfa and was stably transmitted to the offspring. Compared to the wild-type plants, the lignin content of T0 and T1 generation plants was reduced by 45.77% and 31.97%, respectively; there were no significant differences in height and weight of T0 and T1 plants, compared to the wild-type plants. However, the transgenic plant differed from the wild-type plant by softer stems and leaves, larger leaves, fewer flowers and a fewer seeds. The T0 line was susceptible to disease infection, but significantly improved in the second year. The results suggest that the 4CL gene from Amorpha fruticosa can be used to regulate lignin biosynthesis in transgenic forage crops.