*2.4. Quantitative Real-Time PCR (qRT-PCR) Analysis*

To verify the reliability of RNA-Seq data, total RNA was extracted from the leaves and roots of biological triplicates from the two sequenced genotypes, and the template cDNA samples were prepared using an iScrip First Strand Synthesis System Kit (Bio-Rad Laboratories, Hercules, CA, USA). Primers for each PCR reaction were designed to have a melting temperature of 58–62 ◦C and to produce a PCR product between 100 and 200 bp (Table S1). The pepper actin gene (*AY572427*) was used as an internal control. qRT-PCR reactions were performed using a CFX96TMReal-Time System (Bio-Rad Laboratories, Hercules, CA, USA). The reaction conditions were as follows: 95 ◦C for 30 s, 40 cycles of 95 ◦C for 5 s, and 60 ◦C for 30 s. The 2−ΔΔCt method was used to calculate the expression levels of genes.
