## 2 Results
### Analysis of Conserved Motifs and Gene Structure  
Since analysis of gene structure will facilitate an understanding of gene evolution and possible roles, the structure of _DoTPS_ genes in _D. officinal_ was investigated (Figure 2A). The amount of exons ranged from 2 to 14, with an average of 6.6 for all _DoTPS_ genes. _DoTPS32_ contained the most exons (14), whereas _DoTPS8_ and _DoTPS12_ harbored the fewest exons (2). The majority of _DoTPS_ genes (52.9%) had seven exons. Apart from _DoTPS7_, -20_, -24, -27, and -28, most of the genes that clustered in the same group generally possessed a similar exon-intron structure, especially in terms of intron number and exon length (Figure 2A). This conserved exon-intron structure within each cluster was in agreement with the classification of _DoTPS_ genes in a neighbor-joining (NJ) phylogenetic tree based on DoTPS sequences.  
To further elucidate the structural and functional features of DoTPS, 20 conserved motifs of the DoTPS proteins were identified using MEME software (Figure 2B). The lengths of these motifs ranged from 15 to 47 amino acids (Figure S1; Table S5). DoTPS3 contained the most motifs (18/20) while DoTPS31 had only two motifs. Motif 6 was found in all DoTPS proteins, except DoTPS12. Motifs 5 and 10 were the second most common DoTPS proteins (32/34), followed by motifs 1 and 2 (31/34). DoTPS1, -8, and -31 did not contain the DDxxD motif (motif 1), and DoTPS1, -12, and -31 did not contain the R(R)X\({}_{8}\)W (motif 2) motif. Intriguingly, motif 14 was found in the cluster containing DoTPS5, -6, -7, -9, 13, -15, -16, -17, -22, -26, and -29. Motifs 17 and 20 were particularly abundant in the group containing DoTPS2, -3,-10, -18, -19, -20, -21, -23, -25, -27, -28, -30, -32, and -34. Motif 20 only existed in the cluster that included DoTPS4, -14, and -32. Motif 18 was only observed in a small branch that harbored DoTPS2, -3, -24, and -27 (Figure 2B). Despite the different types of motifs among clusters, DoTPS proteins within the same cluster generally possessed similar motifs. The diversity of DoTPS phylogenetic grouping patterns was likely influenced by the gene structure and the location of motifs.  
\begin{table}
\begin{tabular}{l l l l l l l l l l} \hline \hline \multirow{2}{*}{**Name**} & \multirow{2}{*}{**Gene ID 1**} & \multicolumn{2}{c}{**ORF 2**} & \multicolumn{1}{c}{**AA 3**} & \multicolumn{1}{c}{**pl 4**} & \multicolumn{1}{c}{**Mw 5**} & \multicolumn{1}{c}{**AI 6**} & \multicolumn{1}{c}{**II 7**} & \multicolumn{1}{c}{**GRAV 8**} & \multicolumn{1}{c}{**Localization 9**} \\  & & \multicolumn{1}{c}{**(bp)**} & \multicolumn{1}{c}{**(aa)**} & \multicolumn{1}{c}{**pl 4**} & \multicolumn{1}{c}{**(kDa)**} & \multicolumn{1}{c}{**AI 6**} & \multicolumn{1}{c}{**II 7**} & \multicolumn{1}{c}{**GRAV 8**} & \multicolumn{1}{c}{**Localization 9**} \\ \hline _DoTPS10_ & Dxa007746 & 1797 & 598 & 5.73 & 69.73 & 93.61 & 47.41 & \(-\)0.242 & Chloroplast \({}^{\text{\text{\text{\text{\text{\text{\text{\text{\text{\text{\text{\text{\text{ \text{ \texttexttexttexttexttexttexttexttext  
Figure 2: Phylogenetic relationships, exon–intron structure, and protein domain analysis of _DoTPS_ genes in _D. officinal._ (**A**) Phylogenetic relationships and exon–intron structure of _DoTPS_ genes. Exon–intron distribution was performed using GSDS 2.0 server ([[http://gsds.cbi.pku.edu.cn/](http://gsds.cbi.pku.edu.cn/)]([http://gsds.cbi.pku.edu.cn/](http://gsds.cbi.pku.edu.cn/))). Yellow boxes indicate exons, black lines indicate introns. Blue boxes represent upstream/downstream-untranslated regions. (**B**) Phylogenetic relationships and motif structures of _DoTPS_ genes. Phylogenetic tree was generated with MEGA 7.0 using the NJ method. Twenty classical motifs in DoTPS proteins were analyzed by the MEME tool. The width of each motif ranged from 15 to 47 amino acids. Different color blocks represented different motifs.