Sybr premix ex taqtm rt pcr kit

To validate the reliability of expression profiles in RNA-seq data (N₀ VS N₂₂₅), 10 genes were selected for quantitative real-time PCR (qRT-PCR) analyses, using a SYBR premix Ex Taq^tm RT-PCR kit (Takara, Dalian, China). Primers were designed by Primer Premier 5.0 (Premier Biosoft, Palo Alto, Canada) and listed in Supplementary Table S9. GAPDH (Genbank accession number KU246046.1), TUBA-2A (DQ435659.1), and TEF1 (M90077.1) were used as reference. The relative expression value was calculated by the ∆∆Ct method.

To study the expression patterns of the ZmSnRK2 genes, we obtained the expression level of each ZmSnRK2 gene in different tissues by analyzing the released transcription data [29 (link)], and then submitted it to the MeV_4_9_0 [61 (link)] for stratified clustering (Hierarchical Clustering, HCL). Further, different tissues of B73 were collected for RNA extract and qRT-PCR analysis, including root, stem, leaf, filament, anther at the same stage (Heading stage). Besides, the development kernels were also collected, including kernels at 2–30 days after pollination (DAP), endosperm and embryo at 15 DAP. The treatment to vitro kernel was according to the paper described by [30 (link)]. The RNA extraction of materials was followed the protocol of Trizol (TIANGEN, Beijing, China) reagent. RNA was reverse transcribed using the PrimeScript™ RT Reagent Kit with genomic DNA Eraser (Takara, Dalian, China), and quantitative RT-PCRs were performed using a SYBR premix Ex Taqtm RT-PCR kit (Takara, Dalian, China). All the experiments were performed following the manufacturer’s instructions. The primers used for qRT-PCR were listed in Additional file 1: Table S2.