Abstract:0.1% colchicine solution was used dipping on growing point of the seedings of Stevia rebaudiana Bertoni to induce the multiploid (tetraploid) of the plant.If dropping it eight times,the mutagen rates was up to 31.25%.The chromosome number in the tetraploid of S.rebaudiana was n=22 or 2n=44.And the chromosome number in the diploid was n=11 or 2n=22.Observation of the morphology and anatomy of tetraploid showed that the stem of tetraploid was shorter and thicker than the diploid‘s.The tetraploid leaf was much bigger than that of diploid.Leaf length and width was 2.1 times longer, 2.3 times wider, respectively.Leaf thickness was 1.7 times thicker,than that of diploid and the color of the leaf was much deeper.Stoma numbers of tetraploid were reduced, but stoma was larger.The test of the stevioside content showed that the percentage of stevioside content in the leaves of was 15.7%,but diploid was 10.8%.