Site-year comparison of transcript expression. Transcript expression of three selected genes found to be downregulated in all five hybrids by the BRB-seq analysis in 2020. Expression levels were measured by RT-qPCR from the same dataset of 2020 as the BRB-seq, and from a replicate experiment in 2021. RT-qPCR expression is normalized to the Z. mays Ubiquitin2 housekeeping gene (Zm00001d053838). Error bars represent the mean ± 1 standard deviation, n = 3 (RT-qPCR) or n = 4 (BRB-seq). Asterisks indicate unpaired t-test values between high and low N treatments; *P < 0.1, **P < 0.01, ***P < 0.001.