Starch accumulation is important during com development. ADP-glucose pyrophosphorylase (AGPase) is the rate-limiting enzyme in starch synthesis. AGPL is the large subunit of AGPase. Here, we isolated and characterized the large subunit of AGPase gene GhAGPL1 in gladiolus (Gladiolus hybridus). GhAGPL1 was highly expressed in sink organs (cormels and corms). The expression of GhAGPL1 was induced by glucose, sucrose, and mannitol, and it was repressed by abscisic acid (ABA). Overexpression of GhAGPL1 in the arabidopsis (Arabidopsis thaliana) apl1 mutant resulted in complementation of AGPase activity and thus starch synthesis. Silencing GhAGPL1 in gladiolus decreased the transcript level of GhAGPL1 and GhSus, and resulted in the reduction of AGPase activity and starch content in gladiolus corm and cormel. Meanwhile, sucrose content was higher in GhAGPL1-silenced corm. Surprisingly, silencing GhAGPL1 in gladiolus produced smaller corms and fewer number of cormels. Overall, GhAGPL1 contributed to the quality and quantity of gladiolus corms and cormels.