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Figure 1. 

Multiple sequence alignment of the 31 SmDof proteins. Different colors represent identical and conserved amino acid residues, and the red box shows the conserved zinc-finger domain.

Figure 2. 

Evolutionary relationship of SmDof proteins in S. miltiorrhiza and Arabidopsis. Varied colors represent different groups. There were five groups, Groups I−V, with the green circles representing the SmDof proteins of Arabidopsis and the orange circles representing the Dof proteins of S. miltiorrhiza.

Figure 3. 

Phylogeny, conserved motifs and gene structure of SmDof proteins in S. miltiorrhiza. (a) SmDof proteins evolutionary tree. (b) Conserved motifs of the 31 SmDof proteins. Different colors represent 15 different motifs, and the bottom line represents the length of the sequence. (c) Exon/intron structures of SmDofs. Green represents UTRs and yellow represents CDS.

Figure 4. 

Cis-acting elements of SmDof promoters in S. miltiorrhiza. Dof family cis-acting element of S. miltiorrhiza. Different colors represent different classes of cis-acting elements and motifs. Green represents cis-acting elements associated with light, yellow represents cis-acting elements associated with plant hormones, purple represents cis-acting elements associated with cell development, and blue represents environmental stress.

Figure 5. 

Expression profiles of SmDof genes and synthetase genes involved in tanshinones and phenolic acids biosynthesis pathway. (a) Contents of tanshinones and phenolic acids in different tissues of S. miltiorrhiza. (b) Expression profiles of SmDof gene in various tissues of S. miltiorrhiza. (c) Expression profile of SmDof genes under the treatment of ABA induction based on the transcriptome dataset. Red and blue boxes indicate high and low expression levels of SmDofs, respectively. (d) Expression profiles of synthetase genes involved in tanshinones and phenolic acids biosynthesis pathway under the induction of exogenous ABA detected by qRT-PCR. Three biological replicates were performed and the mean ± SD was taken, SD was represented by the error line. * indicates significant difference in t-test (*p < 0.05).

Figure 6. 

Co-expression analysis of SmDof genes and the genes involved in the biosynthetic pathway of phenolic acids and tanshinones. R > 0.5 indicates a positive correlation; R < –0.5 indicates a negative correlation. The data were obtained from the ABA transcriptome dataset.

Figure 7. 

Functional characterization of SmDof12 and SmDof29. (a) Expression patterns of SmDof12 and SmDof29 in four tissues of S. miltiorrhiza. The fold changes of the relative gene expression level in the other three tissues are all normalized to the expression level in flower. (b) Expression patterns of SmDof12 and SmDof29 in hairy roots of S. miltiorrhiza treated with ABA. The fold changes in the relative gene expression level were all normalized to the control expression without induction at the 0 h time point. (c) Subcellular localization of SmDof12 and SmDof29 in tobacco. 35S-YFP is the control group, yellow is the fluorescence of YFP, and blue is the nucleus. Scale bar = 50 μm. (d) Dual-Luc assay showed that SmDof12 could inhibit the activity of SmGGPPS promoter and SmDof29 could promote the activity of SmPAL promoter. Three biological replicates were performed and the mean ± SD was taken, SD was represented by the error line. * indicates significant difference in t-test (* p < 0.05, ** p < 0.01).