|Abstract||Sucrose accumulation is critically important in determining sugarcane productivity. Yet, improvement in sucrose content in sugarcane has been almost stagnant for decades in many sugarcane growing countries, including Australia. This project has made significant improvement in sucrose content in sugarcane varieties by manipulating the activity of a specific gene involved in sucrose metabolism. In a previous study, we created transgenic sugarcane clones with Q117 background, which showed 15-25% improvement in sucrose content under glasshouse conditions. This high-sucrose phenotype was reproduced in the field in this project. The same transgenic technology was successfully applied to Q208A and Q240A and improvement of one to two units in CCS were recorded for seven lines, out of 365 tested, under field conditions. The high-sucrose phenotype was stable across plant and first ratoon crop during the two-year field study. Despite of the increase in sugar content, three lines showed comparable cane yield and fibre content to that of untransformed controls. Molecular analyses of high-sucrose lines confirmed that the phenotype was caused by RNAi-mediated down-regulation of target gene expression. The study demonstrates the potential of targeted gene manipulation for sucrose improvement in already established sugarcane varieties. Considering the growing resistance to genetically modified food crops, use of alternative emerging technologies such as gene editing may be the way forward for realising the same or better results.