2.7. Whole genome expression analysis

Whole genome expression analysis was carried out after culturing murine C3H10T1/2 cells on various samples in triplicate for 12 h. Total RNA was isolated and purified using TRIzol reagent (Invitrogen, Carlsbad, CA, USA). Poly (A) RNA was purified from 1 μg of RNA and fragmented into small pieces. The cleaved RNA fragments were then reverse-transcribed to create cDNA using SuperScript™ II Reverse Transcriptase (cat. 1896649; Invitrogen, USA). U-labeled second-stranded DNAs were then synthesized using the obtained DNA and RNA. Finally, 2 × 150 bp paired-end sequencing (PE150) was conducted on an Illumina Novaseq™ 6000 (LC-Bio Technology Co., Ltd., Hangzhou, China) according to the manufacturer's protocol. Cutadapt software was used to remove reads that contained adaptor contamination. HISAT2 software was used to map reads to the genome and then assembled using StringTie. Then, all transcriptomes from all samples were merged to reconstruct a comprehensive transcriptome using gffcompare software. The expression values were selected with fold change >2 or <0.5 and p value < 0.05 using the R package edgeR, and defined as differentially expressed genes (DEGs). The gene ontology (GO) enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases were analyzed to identify potential pathways.