Validation of selected candidate genes using real time-PCR. We selected ten genes for validation by real time-PCR: CF936190, AF515607, CK089075, CV273041 and CV260219A are the up-regulated genes, and AJ780277, CX183751, CX185631, CV260015 and DN487027 are the down-regulated genes in high photosynthetic rate gene pools. (A) The scatter plot show the relationship between transcript abundance of candidate genes in microarray and the relative expression level using real time-PCR, for which the template was the mixed cDNA of BSA (bulked segregant analysis) pools. (B) The relative expression level using real time-PCR, for which the template was the mixed cDNA of BSA pools and the templates were from the individuals used for constructing BSA pools. Error bars represent the standard deviation. Real time-PCR expression analysis with ANOVA indicated significant differences between the groups with high and low photosynthetic rate (Additional file 7: Table S4), generally consistent with the expression patterns identified by the microarray. The trends are consistent between Real time-PCR expression measured in the gene pool and relative individuals (constructing these gene pools) in general (r = 0.939).