Female B6C3F1 mice (n=4) Furan (0 or 8 mkd) 3 weeks RNA Extraction Agilent 8x60K microarray s: 1- and 2-color arrays Illumina HighSeq RNA-seq: PolyA and ribodepleti on Global gene expression analysis 18hrs 3wks in formalin Paraffin Embedding Frozen - 80°C Divide into three sections Suppl. Fig. S1. Experimental Design. Mice were exposed orally to furan for 21 days (N = 4 per group). Liver was sectioned into three, with one section preserved by flash freezing, the second preserved in formalin for 18 hours followed by paraffin embedding, the third preserved in formalin for 3 weeks prior to paraffin embedding. RNA was extracted from each of these liver samples and every sample was subject to analysis by: (1)
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FemaleB6C3F1 mice
(n=4)Furan
(0 or 8 mkd)3 weeks
RNA Extraction
Agilent 8x60K microarrays: 1-
and 2-color arrays
Illumina HighSeq RNA-seq: PolyA
and ribodepletion
Global gene expression analysis
18hrs 3wksin formalin
Paraffin Embedding
Frozen- 80°C
Divide into three
sections
Suppl. Fig. S1. Experimental Design. Mice were exposed orally to furan for 21 days (N = 4 per group). Liver was sectioned into three, with one section preserved by flash freezing, the second preserved in formalin for 18 hours followed by paraffin embedding, the third preserved in formalin for 3 weeks prior to paraffin embedding. RNA was extracted from each of these liver samples and every sample was subject to analysis by: (1) one-color DNA microarrays; (2) two-color DNA microarrays; (3) poly-A RNA-sequencing; or (4) ribo-depletion RNA-sequencing.
Suppl. Fig. S2. Pathway analysis for (A) ribo-depletion and (B) polyA-enrichment, and (C) 1-colour microarrays, (D) 2-colour microarrays, showing the top 15 enriched pathways for the fresh-frozen samples and enrichment levels of these pathways in the corresponding FFPE samples (pathway significance was calculated in IPA using a Fisher Exact test, -log(p value)=1.3 corresponds to a p=0.05). Venn diagrams show the overlap of the top 15 pathways in each fresh-frozen, 18hr FFPE, and 3wk FFPE. For all analyses, all (i.e.: unmapped) differentially expressed genes with FDR p < 0.05, fold change > +/-1.5 were used.
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-log(
p va
lue)
IPA pathways (Ribo-, unmapped)
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-log(
p va
lue)
IPA pathways (PolyA, unmapped)
A
B
Suppl. Fig. S2. Pathway analysis for (A) ribo-depletion and (B) polyA-enrichment, and (C) 1-colour microarrays, (D) 2-colour microarrays, showing the top 15 enriched pathways for the fresh-frozen samples and enrichment levels of these pathways in the corresponding FFPE samples (pathway significance was calculated in IPA using a Fisher Exact test, -log(p value)=1.3 corresponds to a p=0.05). Venn diagrams show the overlap of the top 15 pathways in each fresh-frozen, 18hr FFPE, and 3wk FFPE. For all analyses, all (i.e.: unmapped) differentially expressed genes with FDR p < 0.05, fold change > +/-1.5 were used.
