Fig. 4

Genomic DNA alters the expression of low-abundance transcripts and leads to false results in Ribo-Zero. a) Genomic DNA significantly altered the quantitation of gene expression levels in Ribo-Zero. The bar plot shows the number of DEGs in Ribo-Zero at different concentrations of contaminating gDNA. The “Correlated” DEGs were considered genes with altered expression levels caused by gDNA contamination, and the “Not Correlated” DEGs were considered genes with altered levels caused by gDNA and/or background noise. The DEGs were detected by comparing libraries with > 0% (Treatment) and 0% (Control) gDNA. The x-axis represents different treatments; the y-axis represents the number of DEGs in each comparison (t test, two-sided, p < 0.05 and |log2(fold-change)|> 1). The red and gray bars represent “Correlated” and “Not Correlated” DEGs, respectively. b) The “Correlated” and “Not Correlated” DEGs were expressed at low levels in the Treatment and Control. Most “Correlated” and “Not Correlated” DEGs in Treatment and Control showed expression levels < 0. The distribution of expression levels of “Correlated” DEGs between libraries with 0.1% and 0% gDNA contamination is not displayed, because only one “Correlated” DEG was detected. The x-axis represents the expression value (log2[FPKM]); the y-axis represents density. The blue line represents Control, the red line represents Treatment. c) “Correlated” and “Not Correlated” DEGs give “false” enrichment results. The plot shows the number of enriched KEGG pathways of DEGs between Treatment and Control in Ribo-Zero. The x-axis represents different treatments; the y-axis represents the number of enriched pathways. The red, gray, and blue bars represent “Correlated”, “Not Correlated,” and all DEG-enriched pathways, respectively