Triploid breeding is a central way to improve growth traits, timber quality, and stress resistance in Populus. In the present study, the morphology and viability of colchicine-induced 2n pollen, triploid production by crossing induced 2n pollen, and identification of genetic constitution of colchicine-induced 2n pollen were conducted in Populus canescens based on optimizing technology for inducing chromosome doubling in pollen. We found that the meiotic stage, injection time, and the interaction between the meiotic stage and injection time had highly significant effects on the 2n pollen production rate. The most effective treatment for inducing 2n pollen was to give 11 injections of 0.5% colchicine solution when pollen mother cells (PMCs) were at the pachytene stage. The highest 2n pollen production rate was 30.27 ± 8.69%. Colchicine occasionally affected ectexine deposition, and some narrow furrows were detected in the ectexine structure. However, no significant difference was observed in the pollen germination rate between natural 2n pollen and colchicine-induced 2n pollen. Moreover, 5 triploids derived from FDR-type 2n pollen were generated by crossing induced 2n pollen, suggesting that colchicine does not eliminate the function of colchicine-induced 2n pollen. However, slower growth of 2n pollen tubes was responsible for a lower triploid production rate.