A click chemistry assay based on a newly discovered DNAzyme, CLICK-17, with azide modified gold nanoparticles (azide-AuNPs) and alkyne capped dsDNA (alkyne-linker DNA) was employed for novel and selective detection of Cu2+ visually. The strategy involved using CLICK-17 to mediate a catalytic reaction for triazole formation between azide-AuNPs and alkyne-linker DNA under the help of Cu2+ (without sodium ascorbate) or Cu+, which eventually led to the aggregation of AuNPs. The obvious color change from ruby red to bluish purple was then observed by the naked eye and the absorbance peak shifted from 525 to 570 nm. Interestingly, CLICK-17 and Cu+-catalyzed click reaction had the best performance compared to either Cu+ alone or CLICK-17 and Cu2+-mediated reaction in terms of the reaction time and sensitivity. This system has been demonstrated to allow quantitative measurement of Cu2+ with a detection limit as low as 26.8 nM and also has high specificity that can distinguish Cu2+ from other metal ions. Further, the method was tested with a real mineral water sample for Cu2+ concentration determination. Satisfactory recoveries of 90.8% and 99.8% were achieved.