The expression profiles of carotenoid metabolism pathway genes are one of the important factors that regulate carotenoid content and composition in the fruit of Citrus cultivars. To identify regulatory factors of gene expressions in the carotenoid pathway, expression quantitative trait loci (eQTL) analysis was applied using a hybrid population. In the analyzed population, significant eQTLs for phytoene synthase (PSY), phytoene desaturase (PDS), ζ-carotene desaturase (ZDS), β-ring hydroxylase (HYb), and zeaxanthin epoxidase (ZEP) were detected. Among these eQTLs, eQTLs for HYb and ZEP were located on their responsible gene loci, respectively. This result indicates that the expression level of HYb and ZEP was influenced by cis-elements in their up-stream regions. Conversely, eQTLs for PDS and ZDS were located in different loci from those of the responsible genes, indicating that the loci were trans-regulating factors. It also suggested that expression levels of PDS and ZDS were regulated by a common transcription factor because their eQTLs were co-localized. Significant eQTL of lycopene β-cyclase (LCYb) and 9-cis-epoxycarotenoid dioxygenase (NCED) were not detected in this population. The major factors to control gene expression depend on each carotenoid metabolism gene, and their combination might cause complex carotenoid metabolism and extend the variation of content and composition in citrus varieties. eQTL analysis was demonstrated as a powerful tool to identify cis- or trans-regulation for carotenoid metabolism genes, as well as in generating functionally defined markers for marker-assisted selection to increase the carotenoid content of citrus in breeding programs.