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The effects of Timentin and cefotaxime (Cef) on shoot regeneration of the London plane tree (Platanus acerifolia Willd.) and their use for the suppression of Agrobacterium tumefaciens in Agrobacterium-mediated genetic transformation were compared.Shoot regeneration was significantly reduced on the media with Cef at concentrations from 100 to 500 mg·L-1. Timentin showed negative effect on plant regeneration at concentrations of 100 and 500 mg·L-1; however, 300 mg·L-1 Timentin was shown to facilitate shoot regeneration significantly and the regeneration frequency increased from 64% (control) to 88%. Effective suppression of A.tumefaciens could be obtained with 500 mg·L-1 Cef, but plant regeneration was completely inhibited at this level. The A. tumefaciens on infected P. acerifolia leaf tissues was visually undetectable after three subcultures on a medium with 300 mg·L-1 Timentin. Considering the effect of Cef and Timentin on plant regeneration and suppression of Agrobacteria, Timentin at 300 mg·L-1 is the preferred application in A. tumefaciens-mediated transformation of P.acerifolia.