To clarify the effects of shading by bagging on the carotenoid accumulation in peach fruit flesh, the fruits of two yellow-fleshed peach cultivars (3D-8 and C18) were analyzed after a bagging treatment (yellow-black double-layered bag) at 50 days after full bloom. The bagging treatment significantly decreased the carotenoid content. The reason for this decrease may be that dark conditions due to bagging upregulates PpPIF3 expression. The resulting increase in PpPIF3 expression level negatively regulates structural genes (PpPSY2, PpPDS, PpZDS, PpLCYE, PpCYB1, PpHYB, PpHYE) expression. Bagging promoted the increase of zeaxanthin and the decrease of xanthophyll, β-cryptoxanthin, α-carotene, β-carotene, and finally reduced the content of total carotenoids. However, bagging minimally influenced the expression of genes related to carotenoid cleavage. Additionally, bagging decreased peach fruit quality and the total phenol content, but increased the reactive oxygen species content.