Fig 1.
Rhizome enlargement process and contents of endogenous hormones during ginger rhizome enlargement.
A and B. Rhizome enlargement process and phenotype of three different stages, including stage 1 (S1), the initial enlargement stage; stage 2 (S2), the middle enlargement stage; and S3, the peak enlargement stage of rhizome development. Bar = 15 cm; The red arrow is the graphical representation of samples. C. Representative hormone production at the three different stages. Three biological replicates of every enlargement stage were performed. Values represent the average ± SD. Different letters indicate significant difference at P = 0.05 level.
Fig 2.
Cluster heatmap and numbers of potential differential unigenes in different development stages of ginger.
A) Cluster heatmap, the different colours are the r2 values of the Pearson’s correlation coefficient; red represents high correlation, and yellow represents low correlation. B) Number of differentially expressed genes (DEGs) in ginger rhizomes between stage 2 (S2) and S1, between S3 and S2, and between S3 and S1; black represents significantly upregulated genes, grey represents significantly downregulated genes. C) Venn diagram of differential unigenes; potential differential unigene numbers between S2 and S1, between S3 and S2, and between S3 and S1.
Fig 3.
Cluster and STEM analysis of differentially expressed genes (DEGs).
Sixteen clusters were obtained using the STEM software. The coloured clusters represent a significant level (p ≤ 0.05). The number at the top is the cluster number. The number at the bottom is the gene number assigned to each cluster.
Fig 4.
Metabolic processes and cellular component activated or repressed at different time points during ginger rhizome enlargement.
The overrepresented Gene Ontology (GO) terms for the combined clusters of genes either upregulated (clusters 8 and 13) or downregulated (clusters 0 and 7) during rhizome enlargement were enriched using BiNGO [28, 30]. Significantly upregulated or downregulated GO terms at each time point during rhizome enlargement are indicated.
Fig 5.
Display of gene expression involved in hormone biosynthesis and signalling pathway.
Significantly differentially expressed genes (DEGs) (log2 fold change (FC) ≥ 1, FDR ≤ 0.05) were visualised using the Mapman software and organised into functional categories (BINs). Blue indicates a decrease and yellow indicates an increase in gene expression (see colour set scale in top right corner). IAA, auxin; ABA, abscisic acid; BA, brassinosteroid; SA, salicylic acid; GA, gibberellin. Detailed information on each gene and its expression level is listed in S5 Table.
Fig 6.
Display of gene expression of transcription factors (TFs).
Significantly differentially expressed genes (DEGs) (log2 fold change (FC) ≥ 1, FDR ≤ 0.05) were visualised using the Mapman software and organised into functional categories (BINs). Blue indicates a decrease and yellow indicates an increase in gene expression (see colour set scale in top right corner). Detailed information on each gene and its expression level is listed in S6 Table.
Fig 7.
Validation of RNA-seq by qPCR.
Stage 1 (S1), S2, and S3 represent the different enlargement stages of ginger rhizome development, respectively. The results represent the mean of three biological replicates with standard deviations. The solid black lines represent the RNA-seq results, and the dashed lines and white circles represent the qPCR results. R2 is the correlation coefficient between RNA-seq and qPCR.
Fig 8.
Connection network between hormones, rhizome diameter, and transcripts.
The networks were visualised using the Cytoscape software (v. 3.7.2). The connection network between transcripts, including hormone synthesis and signalling-related genes, transcription factors, cell division and expansion-related genes, and hormones or rhizome diameter (RD), respectively. Symbol IDs are listed in S3–S5 Tables.
Table 1.
Differentially expressed genes (DEGs) correlated with rhizome diameter in Zingiber officinale Roscoe ‘RY20’.
Fig 9.
Expression profifiling of 30 related genes in 15 different ginger varieties.
Expression analysis of 30 genes at peak enlargement stage S3 (almost grow to mature size) of ginger rhizomes were determined by qPCR and illustrated by heatmap. The Min-Max method was used to standardize the relative expression data of each gene, and the transformation formula: (n-minimum value)/(maximum value-minimum value) was used to make all trait values in a specific interval of 0–1, where n is independent variables.
Fig 10.
Possible positive regulatory network in ginger rhizome enlargement process.