Table 1.

RNA-seq bioinformatic workflow for calling differentially expressed genes.

STEP	TOOLS	CHALLENGES
1. Remove low-quality reads, barcodes, and adapters	Fastx-toolkit, FLEXBAR, or Trimmomatic	Follow recommended protocol
2. Remove mitochondrial and ribosomal sequences	Bowtie2	Sequences from the same or related species should be used
3. Align to reference genome	TopHat2	Incomplete or nonexistent reference genome
4. Call differentially expressed genes	DESeq2, edgeR, or limma	Incomplete or nonexistent reference genome annotation