Gene/genome dosage balance is an essential evolutionary mechanism for organisms to ensure a normal function, but the underlying causes of dosage-imbalance regulation remains poorly understood. Herein, the serial Brassica hybrids/polyploids with different dosages of A and C subgenomes were synthesized to investigate the effects of genome dosages on gene expressions and interaction using RNA-Seq. The expression changes of A- and C-subgenome genes were in accordance with dosage alterations. Dosage-dependent and -independent genes were grouped according to the correlations between dosage variations and gene expressions. Expression levels of dosage-dependent genes were strongly correlated with dosage changes that mainly contributed to dosage effects, while those of dosage-independent genes gave weak correlations with dosage variations that mostly facilitated dosage compensation. More proteinâ€“protein interactions were detected for dosage-independent genes than dosage-dependent ones, as predicted by the dosage balance hypothesis. Dosage-dependent genes more likely impacted the expressions by trans effects, whereas dosage-independent genes preferred to play by cis effects. Furthermore, dosage-dependent genes were mainly associated with the basic biological processes to maintain the stability of the growth and development, while dosage-independent genes were more enriched in the stress response related processes to accelerate adaptation. The present comprehensive analysis of gene expression dependent/independent on dosage alterations in Brassica polyploids provided new insights into gene/genome dosage-imbalance regulation of gene expression. Overall design: The newly expanded third leaves without petioles from six plants of each genotype were collected and immediately stored in liquid nitrogen for RNA extraction. Six samples with two replicates were sequenced via Illumina HiSeqTM 2000.