Quantitative Real-Time PCR
The expression of Sh2 was measured in the fifth leaf, three days after emergence. The first, third and eighth centimeter from the leaf base were harvested in sh2 as meristem, elongation and mature tissues, respectively, based on the cell length profile. In the wild type, these zones were harvested as the first, fifth and tenth centimeter from the leaf base, respectively. Total RNA was diluted to the same concentration (0.4 μg. μL-1) for all samples and first strand cDNA synthesis was performed with superscript II Reverse Transcriptase according to the manufacturer’s protocol (Thermo Fisher Scientific, USA). Quantitative Real-Time PCR was performed with the Takyon qPCR Kits for SYBR Assay (Eurogentec, Belgium) using the UBCP gene (Zm00001d001913) as a housekeeping gene (Manoli et al. , 2012). The primers 5’-GTAAGGGCATCCAAGAGG-3’ and 5’ -AAGCGGCTCTT ACCATAC-3’ were designed to amplify the Sh2 gene.