## Results
### Gene Expression Analysis across Different Tissues  
In order to investigate the DEGs among the four _E. rituximab_ tissue samples, the FPKM method was employed, and the unigenes with a significant level of FPKM value above 0.3 were selected. A total of 54,441 unigenes from the root, 60,451 from the stem, 70,894 from the leaf, and 47,097 from the latex were found to have higher FPKM values above 0.3 (Table S2). Among them, 29,853 unigenes were expressed in all of the four tissues, while 3392, 1932, 6890, and 4191 unigenes were specifically expressed in the root, stem, leaf, and latex, respectively (Figure 3A). The pairwise comparison of DEGs between latex and three other tissues revealed that the numbers of DEGs between latex and leaf, latex and root, and latex and stem were 5927, 5827, and 5692, respectively. Among all the \begin{table}
\begin{tabular}{c c c} \hline \hline
**Database** & **Number of Unigenes** & **Percentage (\%) b** \\ \hline Nr a & 46,804 & 51.08 \\ SwissProt & 36,923 & 40.30 \\ KOG & 13,292 & 14.50 \\ GO & 38,652 & 42.18 \\ KEGG & 17,447 & 19.04 \\ Annotated in all databases & 7835 & 8.55 \\ Annotated in at least one database & 52,448 & 57.24 \\ \hline \end{tabular}  
* Nr: NCBI non-redundant protein database; b Percentage of annotated unigenes in the total 91,619 assembled unigenes. KOG: EuKaryotic Orthologous Groups (database); GO: Gene Ontology (database); KEGG: Kyoto Encyclopedia of Genes and Genomes.  
\end{table} 
Table 2: Statistics of annotations for assembled unigenes in the transcriptome of _E. rituximab_. DEGs, 2922 of them were found to be overlapped across all three sets of pairwise comparisons as shown in the Venn diagram (Figure 3B). Moreover, the cluster analysis of these DEGs revealed that the gene expression pattern in latex was distinctly different from that in the other three tissues, suggesting that the transcriptional regulations of DEGs for their functions were unique in the latex (Figure 3C). The DEGs were further applied for the GO enrichment analysis and KEGG pathway enrichment analysis (Table 3). Based on the KEGG analysis, a large portion of DEGs, which exhibited higher expression levels in the latex compared to the other tissues, were found to be associated with several secondary metabolite pathways, such as, the \"Sesquiterpenoid and triterpenoid biosynthesis\" (ko00909), \"Terpenoid backbone biosynthesis\" (ko00900), and \"Steroid biosynthesis\" (ko00100). On the other hand, the unigenes relevant to photosynthesis were significantly down-regulated in latex compared to the leaf and stem tissues, which could be explained as latex is definitely not a structure required for photosynthesis. Figure 2: Classification of the function of the _E. rituximab_ unigenes based on the KOG (**A**) and GO (**B**) analysis. Figure 3: Expression analysis of the _E. rituximab_ unigenes. (**A**) Venn diagram analysis for the unigenes with their Fragments Per Kilobase Million (FPKM) values above 0.3 in different tissues; (**B**) Differentially Expressed Genes (DEG) analysis between tissues: latex vs. root, latex vs. stem and latex vs. leaf; (**C**) Heatmap analysis showing the expression pattern of DEGs in different tissues. \begin{table}
\begin{tabular}{c c c c c c} \hline
**Pathway Term** & **Number** & \begin{tabular}{c} **Pathway Term** \\ **Late vs. Leaf** \\ \end{tabular}  & **Number** & \ \begin{tabular}{c} **Pathway Term** \\ **Late vs. Root** \\ \end{tabular}  & **Number** \\ \hline \multirow{7}{*}{
\begin{tabular}{c} \  
To verify the reliability of DEGs identified by the FPKM method, 18 DEGs involved in the sterol biosynthesis were selected and their gene expression levels in the root, stem and leaf tissues were monitored by RT-qPCR analysis. As shown in Figure S6, the gene expression patterns for 15 DEGs (except for _MVD_, _HDR_, and _EtOSC2_) matched the results calculated by the FPKM method based on the transcriptome, suggesting that the FPKM method was reliable for gene expression analysis.