Abstract: In order to enhance the resistance to fungi diseases of K 326, a cultivar of flue-cured tobacco, a chitinase chiA gene from Serratia marcescens was introduced into tobacco by an agro-bacterium mediating method using tobacco lamina of regenerated plant as an accepter and transgenic plants with kanamycin-resistance were obtained.The PCR examination confirmed that the chitinase chiA gene was incorporated in tobacco genome.Meanwhile, the anti-Kan level of tobacco, the effect of pre-culture time of accepter on transformation and anti-Kan shoots rooting were studied.The results showed that kanamycin at the level of 50mg/L could restrain tobacco lamina from regeneration completely, the highest transformation rate was got when the lamina was pre-cultured for 2 days and rooting rate of the transformed shoots cultured in MS containing 0.2mg/L IAA was increased remarkably.