RNA-seq analysis
Total RNA was extracted from the back skin lesions of the IMQ-treated and high-dose allicin-treated groups for transcriptome sequencing. RNA was isolated and purified using Trizol reagent (Invitrogen, Carlsbad, CA, USA). The RNA amount and purity of each sample were quantified using NanoDrop ND-1000 (Wilmington, DE, USA). The final cDNA was sent for library preparation and sequencing on an Illumina Novaseq™ 6000 at LC Bio Technology (Hangzhou, China). With StringTie and Ballgown, we estimate the expression levels for all transcripts and calculate the expression levels for mRNAs with FPKM (FPKM = [total exon fragments/mapped reads (millions) × exon length (kB)]). The differentially expressed genes (DEGs) were selected by R package edgeR or DESeq2 with fold change > 2 or fold change < 0.5 and p-value < 0.05. Following that, we conducted gene ontology (GO) enrichment and Kyoto encyclopedia of genes and genomes (KEGG) enrichment analyses of the differentially expressed mRNAs in the two groups.