*2.6. Gene Expression Analysis*

Quantitative real-time PCR (qRT-PCR) was conducted to test the expression differences of candidate genes between accessions with extreme mesocotyl length (Table S4). The mesocotyl part was sampled at the 52 h after germination before the coleoptile excavated. Total RNA was extracted according to the Trizol method, cDNA was synthesized with the HiScript II 1st Strand cDNA Synthesis Kit (Vazyme, China). Primers (Table S5) were designed with the primer premier 5.0 software (http://www.premierbiosoft.com/; last accessed Jan 2019). The PCR procedure was conducted in a volume of 20 μL, containing 2μL cDNA, 0.4 μL of each primer (μM), 10 μL ChamQ Universal SYBR qPCR Master Mix. *OsActin1* was used as the internal control to normalize the expression level of different samples. All assays were performed in two independent experiments, each with three repetitions.
