Quantitative reverse transcription PCR (qRT-PCR) analysis

Total RNA from wheat flag leaves and grains, which were grown under normal conditions during the whole filling stage, was isolated according to the manufacturer’s protocol (flag leaf, Trizol Up, Trans, China; grain, RNAprep Pure Plant Kit, TianGen, China). qRT-PCR was carried out in Quantiative analysis was performed using the Bio Rad CFX Manager system. This method normalizes the expression of a specific gene versus a control reference with the formula 2-ΔΔCT. In this study, the mRNA levels for two stably expressed genes, tubulin and actin, were evaluated as control genes for qRT-PCR analyses. The information from all of the genes in the qRT-PCR experiments are listed in Table 1http://www.sciencedirect.com/science/article/pii/S0885576515000338 - tbl1.