Due to the long flowering cycle, unpredictable flowering period and low seed setting rate of most bamboo plants, bamboo breeding has always been a difficult problem in the research of bamboo plants. Polyploid breeding, as a common means of plant breeding, is able to obtain progeny with excellent traits through artificial induction. In bamboo breeding, there are fewer studies on polyploid breeding. In this study, based on the existing regeneration system of Dendrocalamus asper, the embryonic callus of D.  asper were treated with colchicine using the liquid suspension method and the solid medium mixed culture method, respectively. The results showed that, based on the differentiation and browning rates of the callus, the better results were obtained by treating the callus with 50 mg∙L^–1 colchicine for 48–72 hours using the liquid suspension method. A total of 54 regenerated plants were obtained in the experiment, including 7 control plants, and 16 chromosome doubled plants were successfully obtained from D. asper using flow cytometry to test all the regenerated plants. In terms of chromosomal doubling, treatment with 100 mg∙L^–1 colchicine for 48 h produced the highest number of chromosome-doubled plants with a polyploidy rate of 54.54%. The 12-ploid plants presented larger and thicker leaves, larger lower epidermal stomata, and other traits, implying their superiority in stress tolerance physiology. This study provides an efficient polyploid breeding technique based on the in vitro indirect regeneration system of bamboo, and offers a new solution for breeding new polyploid germplasm of bamboo.