Longan Gene Expression Analysis

The total RNA of longan EC, ICpEC, GE, and EC treated with different hormones were extracted by TransZolUp kit (TransGen Biotech), and the total RNA of different development stages of the zygotic embryo was extracted by BioTeke kit (Cat. No. RP3301). The cDNA was synthesized with a Hifair^®III 1st Strand cDNA Synthesis SuperMix for qPCR (gDNA digester plus) (Yeasen). Primer design was performed using Primer3 online software (https://primer3.ut.ee/cgi-bin/primer3/primer3web_results.cgi) (Supplementary Table 1). DlACTB, DlEF-la, and DlUBQ were used as internal control genes (Lin and Lai, 2010 (link)). The qPCR system was 20 μL, including HRbio™ qPCR SYBR^®Green Master Mix (No Rox) (Heruibio), 8.2 μL ddH₂O, 1 µL of 10-fold diluted cDNA, and 0.4 μL specific primer pairs, with three replicates. The reaction procedure was 95°C for 30 s, followed by 40 cycles of 95°C for 10 s and 58°C for 30 s, and 2^−ΔCT was used to calculate genes expression (Livak and Schmittgen, 2002 (link); Vandesompele et al., 2002 (link)). SPSS software was used for significant analysis of gene differences, and GraphPad 8.0.2 was used for the draft.