RNA-Seq
Total RNA was extracted from tobacco leaf samples and checked for concentration, purity, and integrity. Qualified RNA samples were used for mRNA library construction. Messenger RNA was enriched using poly-T oligo-attached magnetic beads, fragmented, and reverse-transcribed into cDNA. The cDNA fragments were subjected to end repair, A-tailing, adapter ligation, purification, and PCR amplification to generate sequencing libraries. After quality assessment, the qualified libraries were sequenced on an Illumina NovaSeq 6000 platform to generate 150 bp paired-end reads.