1. Introduction
Abiotic stresses including high temperature, drought, cold, and salt have important effects on plant development and growth. Some transcription factors regulate the transcript levels of their target genes under stress by binding to specific DNA sequences in their target promoters [1,2,3]. Therefore, the regulatory networks of various biological processes can be understood by studying plant transcription factors.
The GRAS transcription factor gene family is named after three originally identified members: gibberellic acid insensitive (GAI), GA1 repressor of GA1 (RGA), and scarecrow (SCR) [4,5,6]. Most GRAS proteins contain only one domain, and the C-terminal region of this domain is highly conserved, which is often referred to as the GRAS domain. It contains five units: SAW, leucine heptapeptide repeat I (LHRI), VHIID, PFYRE, and leucine heptapeptide repeat II (LHRII) [7]. However, some GRAS proteins still contain one domain and another functional domain or two domains [7]. The high variability of the N-terminal region of GRAS proteins determines the regulatory proteins’ specificity [8]. The GRAS transcription factor family has been found in many plants, but the family classification varies slightly in different species [9]. For example, in soybean (Glycine max), 117 GRAS genes are divided into 11 subfamilies: AtSCL4/7, Os19, Os4, HAM, DELLA, DLT, AtPAT1, LISCL, AtSCR, AtSCL3, and AtSHR [10]. In tomato (Solanum lycopersicum), 53 GRAS genes have been identified and classified into 13 subfamilies: HAM, LAS, SCL4/7, SCR, SCL9, SCL28, DELLA, SHR, PAT1, Os4, Os19, GRAS37, and Pt20 [11]. In Brachypodium distachyon (Brachypodium distachyon (Linnaeus) P. Beauvois), 63 GRAS genes are classified into 10 subfamilies: HAM, PAT1, SHR, DELLA, SCL3, SCL4/7, LAS, SCR, DLT, and LISCL [12]. In potato (Solanum tuberosum L.), 52 GRAS genes have been identified and classified into 8 subfamilies: DELLA, LAS, HAM, PATI, SCR, LISCL, SHR, and SCL3 [13]. However, in Populus trichocarpa (P. trichocarpa), 93 GRAS genes are divided into 13 subfamilies: Os19, HAM, Os4, Pt20, DLT, AtSCl3, AtSHR, AtPAT1, AtSCR, AtSCL4/7, AtLAS, DELLA, and LISCL [9]. These studies suggest that members of the GRAS gene family differ in different plants. Their classification reflects their evolutionary history.
The diversity of the GRAS gene family is consistent with its high diversity of functions [14]. To date, functional identification of GRAS transcriptional regulators has occurred mainly in Arabidopsis thaliana and rice. This has shown that they have important roles in the regulation of plant development and growth, including multiple growth regulatory signals and environmental signals such as abiotic/biotic stresses, light, and phytohormones [15]. Sun et al. [16] reviewed the major biological functions of the ten subfamilies of GRAS transcriptional regulators, and the results are reported in Table 1.
Although the GRAS gene family has been studied for many years, the mechanisms and evolutionary dynamics of this gene family in woody plants are still not fully understood. Differences in loss and retention of duplicated gene family members between woody and herbaceous species may help in identifying genes with specialized roles in the adaptive evolution of different lineages. The cacao is called “soft gold” because of its high value. The flowers of cacao trees have ornamental value, and cacao is the main ingredient in chocolate and cacao powder [27]. In addition, cacao beans have important uses in the pharmaceutical and cosmetic industries. Cacao is receiving increasing attention for its potential health benefits because it is rich in polyphenols, particularly flavonoids [28]. However, cacao production is hampered for a number of reasons. Therefore, it is of great value to study the cacao tree. The phylogenetic relationship, conserved domain, and collinearity analysis of this family can provide new ideas for further functional analysis. In 2010, Corti et al. [29] carried out the sequencing and assembly study of the T. cacao genome, and since then, researchers have successively identified and analyzed its important gene families, such as the NAC gene family [30], WRKY gene family [31], and the GPX transcription factor family [32]. In 2013, researchers completed an analysis of the metabolome and transcriptome of the cacao tree [33]. Our knowledge about the expansion and diversification of this gene family in plants is presently limited to the herbaceous species Arabidopsis. To date, the GRAS gene family has not been identified and classified in T. cacao.
In this study, we identified 46 GRAS gene family members and conducted a comprehensive genome-wide analysis of the GRAS gene family of the cacao tree, including gene structure, domain analysis, intron/exon, chromosome location, subcellular localization, and cis-acting elements of GRAS genes. In addition, we analyzed the phylogenetic relationship of GRAS proteins between T. cacao and A. thaliana. Furthermore, we performed the gene duplication pattern of T. cacao GRAS proteins, and we analyzed a syntenic analysis of GRAS proteins among T. cacao, A. thaliana, P. trichocarpa, and Sesamum indicum (S. indicum). The results of this study lay the foundation for further studies of the biological function of genes in T. cacao and provide a reference for subsequent molecular mechanisms.
2. Materials and Methods
2.1. Identification of GRAS Gene Family in T. cacao
The genome file, protein file, coding sequences (CDS), and annotation files of T. cacao were obtained from Ensembl Plants (
The HMM model of HMMER (version 3.1b2) was used to screen GRAS protein candidate members of T. cacao twice to determine the final target members. Firstly, the downloaded HMM profile was employed using the HMMER v3.3.2 program to search for proteins containing target GRAS domains as the initial filtering results, and ClustalW (version 2.1) was used to perform multiple sequence alignment for the initial target proteins [36]. Secondly, to expand the filtering scope, we constructed a new HMM model with e-value < 1 × 10−20. The new model was used to filter second target proteins using HMMER (version 3.3.2), with e-value < 0.05. The two results were combined and used as the final candidate proteins. Finally, the NCBI Conserved Domain Search (
2.2. Physicochemical Properties and Subcellular Localization Analyses of TcGRAS Genes
The online software ProtParam (
2.3. Phylogenetic Analysis and Classification of TcGRAS Genes
To provide family classification of GRAS genes and understand their phylogenetic relationships, a rooted neighbor-joining (NJ) phylogenetic tree between T. cacao (TcGRAS) and Arabidopsis GRAS proteins was built using the MEGA 11 software (version 11.0.11) [39,40]. The TcGRAS genes were classified according to their phylogenetic relationship with A.thaliana GRAS members. We obtained Arabidopsis GRAS protein sequences from TAIR (
2.4. Gene Structure and Conserved Motif Analyses of TcGRAS Genes
The conserved motifs of the TcGRAS proteins were predicted by using the online program MEME (
2.5. Chromosomal Mapping and Cis-Acting Regulatory Analyses of TcGRAS Genes
The online program MG2C (
2.6. Gene Duplication and Synteny Analyses of TcGRAS Genes
The ‘MCScanX’ function of the TBtools software with default parameters was used to predict gene duplications of TcGRAS genes. MCScanX Diamond output was used to calculate the replication events of the T. cacao genome. The Duplicate_gene_classifier program in MCScanX (
2.7. GO Annotation Analyses of T. cacao TcGRAS Genes
The DAVID online program was used to annotate TcGRAS genes. The official gene sample lists of TcGRAS genes were uploaded to the program. The analysis included three parts: molecular function, cell components, and biological processes. The R programming language (version 4.1.3) was used to visualize the GO annotation analysis [47].
3. Results
3.1. Identification of GRAS Members in T. cacao
A total of 70 GRAS protein candidates were obtained from the initial filtering. After the second filtering, 53 candidate proteins were obtained. Finally, 46 GRAS genes were identified by redefining conserved domains and deleting repeats (Supplementary File S1). The identified genes were named from TcGRAS1 to TcGRAS46 according to their chromosomal position. The number of amino acids (aa), average molecular weight (MW), theoretical pI, instability index, and aliphatic index of identified TcGRAS genes were statistically analyzed (Table 2). The number of amino acids of the TcGRAS genes ranged from 347 (TcGRAS21) to 1659 (TcGRAS22) aa, and the molecular weight ranged from 39,709.90 to 191,183.51 Da. The results showed that 44 GRAS proteins were acidic with pI values less than 6.5. Two (TcGRAS2 and TcGRAS19) were neutral, with pI between 6.5 and 7.5. The results of the instability index analysis showed that most TcGRAS proteins were unstable, except for TcGRAS8, TcGRAS12, TcGRAS24, and TcGRAS33. Prediction of the subcellular localization of TcGRAS proteins by the online software BUSCA tool revealed that 41 TcGRAS proteins were mainly located in the nucleus, 4 in the chloroplasts, and only 1 in the endomembrane system.
3.2. Phylogenetic Analysis of TcGRAS and AtGRAS
To explore the evolutionary relationship of GRAS proteins between T. cacao and A. thaliana, we performed a multiple sequence alignment of 46 TcGRAS proteins and 34 AtGRAS proteins, and then constructed an unrooted phylogenetic tree using the MEGA 11 software (Figure 1). According to the homology of GRAS proteins in A. thaliana, 46 TcGRAS proteins were divided into 10 clades, which were HAM, LISCL, LAS, SCL4/7, SCR, DLT, SCL3, DELLA, SHR, and PAT1. It is notable that 5 of the 46 TcGRAS proteins were not classified as any of these subfamilies; therefore, we grouped TcGRAS22 as UN1 and TcGRAS11, TcGRAS13, TcGRAS17, and TcGRAS33 as UN2. The largest clade was subgroup LISCL, which contained ten TcGRAS members (TcGRAS8, TcGRAS24, TcGRAS25, TcGRAS27, TcGRAS39, TcGRAS40, TcGRAS41, TcGRAS42, TcGRAS43, and TcGRAS46), whereas subgroups UN1, DLT, SCL4/7, and LAS only had one member. Subgroups UN1 and UN2 only contained T. cacao members, meaning that these genes may have been specialized during the evolutionary process.
3.3. Gene Structure, Conserved Motifs, and Domain Analyses of TcGRAS Genes
To understand the structural diversity and similarity of GRAS gene family members in the cacao tree, we used the Gene Structure View function of the TBtools software to construct a triad map of the evolutionary tree, gene structure, and motif of GRAS gene family members, as shown in Figure 2. We first performed an individual phylogenetic tree using an NJ method consistent with the phylogenetic analysis between TcGRAS and AtGRAS (Figure 2A).
To further understand the characteristics of the GRAS gene families in T. cacao and the conserved motifs shared among different subfamilies, we used the Multiple Expectation Maximization for Motif Elicitation program to find the conserved motifs. A total of 10 conserved motifs were predicted and named Motif 1–10 (Figure 2B and Supplementary File S2). Early sequence analysis indicated that the GRAS proteins typically share a variable N terminus and a highly conserved C terminus. In this study, we found that the C-terminal regions contained a highly conserved domain (Motif 6). Three proteins did not contain this conserved motif, including TcGRAS2, TcGRAS20, and TcGRAS21, and we hypothesized that the C-terminal region of these GRAS proteins was truncated, lacking part of the GRAS domain.
We used the Gene Structure Display Server 2.0 program to construct a domain analysis of TcGRAS proteins (Figure 2C). We found a total of eight types of conserved domains. All the GRAS genes contain the GRAS domain or GRAS superfamily domain. In addition, the domains of GRAS members in DELLA also have the DELLA superfamily. The TcGRAS2 gene has a GRAS superfamily and TB2_DP1_HVA22 superfamily domain. The TcGRAS22 gene has a GRAS superfamily, ZnF_BED, DUF4413, Dimer_Tnp_hAT, and Peptidase_c48 superfamily domain.
3.4. Chromosomal Mapping and Cis-Acting Regulatory Analyses of TcGRAS Genes
The location of TcGRAS genes was obtained from genome annotation files. A total of 46 TcGRAS genes were randomly distributed on 10 chromosomes and were named from TcGRAS1 to TcGRAS46 according to their positions on the chromosomes (Figure 3A). Chr01 and Chr04 had the largest number (nine, 19.57%) of TcGRAS genes, followed by Chr09 with eight members (17.39%). In contrast, Chr05, Chr06, and Chr10 contained only two TcGRAS genes each (4.35%). Chr04 contained seven subgroups of TcGRAS genes, as shown in Figure 3B, while Chr05, Chr06, and Chr10 contained only two subgroups each. Subgroup DLT was only observed on Chr03, subgroup SCL4/7 was only observed on Chr01, and subgroup LAS was only observed on Chr07.
Cis-acting elements regulate transcription initiation and transcription activity by binding to transcription factors. To explore the promoter function of TcGRAS genes, we extracted 2000 bp sequences upstream of the transcription start site. Then, we submitted these to the online program PlantCARE. Seven types of important cis-acting elements were obtained after sorting and screening, including light-responsive element, abscisic acid-responsive element, MeJA-responsive element, defense and stress-responsive element, low-temperature-responsive element, auxin-responsive element and gibberellin-responsive element. The light-responsive element was found in all promoter regions of TcGRAS genes. In addition, more than half of the 46 GRAS genes had the abscisic acid-responsive element, MeJA-responsive element, and gibberellin-responsive element. Compared with the MADS-Box transcription factor family in cacao tree, the GRAS gene family contains significantly more light-responsive elements, abscisic acid-responsive elements, and MeJA-responsive elements. The distribution of these cis-acting elements is shown in Supplementary File S3.
3.5. Gene Duplication and Syntenic Analysis of TcGRAS Genes
Genome-wide analysis of cacao tree gene replication by MCScanX software revealed 2148 tandem duplicated genes in the cacao tree genome, while only 2 pairs of tandem duplicated genes were present among 46 TcGRAS genes (Figure 4). The analysis showed that one pair of tandem duplication genes (TcGRAS24 and TcGRAS25) was located on Chr04, and another pair (TcGRAS42 and TcGRAS43) on Chr09. In addition, the substitution ratio of non-synonymous (Ka) to synonymous (Ks) mutations (Ka/Ks) of two pairs were calculated (Table 3). The Ka/Ks values of both pairs were more than 1, indicating that these genes were positively selected over the course of evolution, and the novel protein functions may be beneficial to the survival and reproduction of T. cacao.
The MCScanX showed that there were 2767 segmental duplications in the genome of T. cacao, and only 6 pairs of fragmental duplicated genes were predicted out of 46 identified TcGRAS genes. The Advanced Circos function of the TBtools software was used to visualize the segmental duplication of GRAS genes on 10 chromosomes, as shown in Figure 4. Chr01 contained three duplicated genes, and Chr02, Chr03, and Chr04 contained two duplicated genes, while Chr05, Chr06, and Chr09 each contained only one duplicated gene. However, Chr07, Chr08, and Chr10 did not contain any segmental duplicated genes.
The syntenic analyses of TcGRAS genes with the GRAS genes of A. thaliana, P. trichocarpa, and S. indicum were separately analyzed to find homologous gene pairs (Figure 5). A total of 36 GRAS genes of T. cacao had a syntenic relationship with the GRAS genes of A. thaliana (16), P. trichocarpa (32), and S. indicum (30). Some genes had multiple syntenic relationships with other closely related species. Therefore, a total number of 25 (Supplementary File S4), 77 (Supplementary File S5), and 48 (Supplementary File S6) GRAS genes of A. thaliana, P. trichocarpa, and S. indicum, respectively, had synteny with 36 GRAS genes. Furthermore, it was found that 13 GRAS genes existed in these 4 plants at the same time (Figure 6). Two homologous GRAS genes existed in T. cacao and S. indicum rather than in P. trichocarpa and A. thaliana. Similarly, T. cacao, P. trichocarpa, and S. indicum had 13 homologous TcGRAS genes that did not exist in A. thaliana, T. cacao, P. trichocarpa, and A. thaliana had 1 homologous TcGRAS gene that did not exist in S. indicum, and T. cacao, S. indicum, and A. thaliana had 2 homologous GRAS genes that did not exist in P. trichocarpa. Five homologous GRAS genes existed in T. cacao and P. trichocarpa but did not exist in A. thaliana and S. indicum.
3.6. GO Annotation of T.cacao TcGRAS Proteins
To understand TcGRAS protein function in different biological processes, we performed a GO annotation analysis of the TcGRAS genes (Figure 7), and the GO numbers are shown in Supplementary File S7. The analysis of the cellular composition showed that most of the TcGRAS proteins were mainly concentrated in the nucleus. The analysis of biological processes showed that the TcGRAS genes were involved in many biological processes. The large portions of GRAS proteins were involved in transcriptional regulation. Otherwise, some TcGRAS proteins were involved in the negative regulation of biological processes, for example, negative regulation of seed germination and the gibberellic acid-mediated signaling pathway. In addition, some TcGRAS genes also respond to abiotic stresses and regulate plant organ development. The analysis of the molecular functions of TcGRAS genes revealed that they had functions in transcription factor activity and sequence-specific DNA binding.
4. Discussion
In this study, we provided the first comprehensive analysis of the GRAS gene family in T. cacao. Based on the latest genome sequences and annotation files, we identified 46 GRAS genes distributed across 10 chromosomes in the T. cacao genome. These 46 TcGRAS genes were classified into 12 subgroups (HAM, LISCL, LAS, SCL4/7, SCR, DLT, SCL3, DELLA, SHR, PAT1, UN1, and UN2) according to their phylogenetic relationship with A. thaliana. We found that the GRAS gene family members were unevenly distributed among subgroups; for instance, the subgroups of UN1 and UN2 only contained T. cacao members, and the member number of subgroups of SCR and SCL3 in T. cacao was more than that in A. thaliana. During the evolution of gene families, the gene structure changes in response to environmental changes to acquire new functions. The structural analysis of TcGRAS genes according to phylogenetic relationships showed that different subgroups had different gene structures and conserved motifs, while the same subgroup had similar motifs and gene structures, which meant that members of the same subgroup had similar functions. Since T. cacao and A. thaliana were exposed to different environments during their evolutionary processes, the number of GRAS genes in their subgroups became different as GRAS genes differentiated.
By analyzing the intron/exon structure of the TcGRAS genes, we found that majority of these genes were free of introns, which was similar to the observed lack of introns in Arabidopsis and rice GRAS genes [8]. A previous study showed that ancestors of each eukaryote had intron-rich genes and that extensive loss and insertion of introns from most genes may have occurred due to selective pressure, with gene duplication accelerating this process [48,49]. Nevertheless, some GRAS genes have evolved different intron/exon structures, indicating that they likely evolved new specialized functions to adapt to their environment.
Tandem and segmental duplications are thought to be the main mechanisms contributing to the expansions of gene families in plants [50]. Both tandemly and segmentally duplicated genes that have been retained in plant genomes play important roles in adaptive responses to environmental stimuli [51,52]. The collinearity analysis in our study showed that there were two pairs of tandem duplication and six pairs of segmental duplication events in the T. cacao GRAS gene family, and this might play an important role in the GRAS family expansion in T. cacao.
The cis-acting elements play a vital role in regulating gene expression during plant growth and development [53]. The promoter analysis showed that the light-responsive element was found in all promoter regions of TcGRAS genes. In addition, more than half of the 46 GRAS genes had the abscisic acid-responsive element, MeJA-responsive element, and gibberellin-responsive element, which made it possible to study the function of these genes in the future.
To further analyze the function of GRAS transcription factors in T. cacao, we studied the end of the genotype affected by functional diversity after GO enrichment analysis, and the results showed that the majority of cacao GRAS proteins play an important role in many different biological processes, including abiotic stresses and plant organ development.
5. Conclusions
In this study, we identified and systematically analyzed the GRAS gene family in T. cacao. Based on the genomic data of the cacao tree, we finally identified 46 GRAS genes using double HMM profiles. These 46 GRAS genes were distributed on 10 chromosomes and phylogenetically divided into 12 subfamilies, with highly similar gene structures and conserved motifs within the same subfamily. Cis-acting element analysis indicated that GRAS genes may be involved in various abiotic stress responses. In addition, we found that tandem and segmental duplications contribute to the expansions of the GRAS gene family. A further syntenic analysis showed that the functions of TcGRAS genes might be speculated from the function of GRAS genes in other plants. Through GO analysis, we found that most of the TcGRAS genes were involved in transcriptional regulation. In summary, the results provide information for further research of the TcGRAS genes’ function and lay the foundation for further investigation.
Conceptualization, S.H.; methodology, S.H.; software, S.H. and Q.Z.; formal analysis, S.H.; investigation, S.H.; resources, S.H. and J.C.; writing—original draft preparation, S.H.; writing—review and editing, Y.G.; visualization, S.H. and J.M.; supervision, S.H., C.W. and J.D.; project administration, Y.G.; funding acquisition, Y.G. All authors have read and agreed to the published version of the manuscript.
Not applicable.
Not applicable.
Data are contained within the article or
We thank Qianqian Zhang (Chinese Institute for Brain Research, College of Biological Sciences and Technology, Beijing Forestry University) and Ang Dong (Center for Computational Biology, College of Biological Sciences and Technology, Beijing Forestry University) for the technical support. This research was supported by the National Natural Science Foundation of China (31370669).
The authors declare no conflict of interest.
Footnotes
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.
Enlarge this image.Figure 1. Phylogenetic tree of GRAS genes in T. cacao and A. thaliana. The GRAS genes in T. cacao and A. thaliana are shown in red and green, respectively. The tree branched the GRAS proteins into different subgroups, illustrated by different colored clusters within the clade. TcGRAS are divided into twelve subgroups according to the subgrouping of A. thaliana. The phylogenetic tree was constructed using the maximum likelihood (ML) method with 1000 bootstrap replications of the MEGA 11 software.
Enlarge this image.Figure 2. Phylogenetic relationship, conserved motifs, and gene structure of TcGRAS genes. (A) An unrooted phylogenetic tree of TcGRAS proteins was constructed using MEGA 11 software under the ML method with 1000 bootstrap replicates. (B) Conserved TcGRAS proteins’ motifs performed by MEME. The colored boxes refer to motifs. The black lines refer to non-conserved sequences. The scale bar is 300 amino acids. (C) The domain analyses of TcGRAS proteins were performed under the Gene Structure Display Server 2.0 program. The domains are displayed in different colored boxes.
Enlarge this image.Figure 3. (A) Physical distribution of TcGRAS genes among 10 chromosomes. (B) Number of TcGRAS subfamilies on each chromosome.
Enlarge this image.Figure 4. Schematic diagram of the duplication patterns of the TcGRAS genes. The red lines show segmental duplications of TcGRAS gene pairs. The gray lines show segmental duplications of all gene pairs in the T. cacao genome. The first ring from outside indicates the chromosomal localization of 46 putative GRAS genes in T. cacao. The second and third rings from outside represent the density of genes on the chromosomes. The blue-to-red scale bar on the right indicates the number of SNPs within 1 Mb window size.
Enlarge this image.Figure 5. Visualization of the syntenic analysis. Synteny of the GRAS genes in T. cacao with the GRAS genes of A. thaliana (A), T. cacao (B), and S. indicum (C) was visualized by MCScanX analysis of TBtools software. Gray lines between the genomes show all synteny blocks, and red lines between the genomes indicate the synteny between the genes.
Enlarge this image.Figure 6. Venn diagram of the identical and different GRAS genes among T. cacao, A. thaliana, P. trichocarpa, and S. indicum.
Enlarge this image.Figure 7. GO annotation of TcGRAS proteins. Green represents biological processes, purple represents cellular components, and orange represents molecular functions.
Recent advances in biological functions of the GRAS transcription factor gene family.
| Gene | Subfamily | Function | Species | Classified Subfamilies | Reference |
|---|---|---|---|---|---|
| HAM | HAM | Maintenance of stems | Petunia hybrida | unclassified | [ |
| LISCL | LISCL | Regulating the transcription process during microsporogenesis | Lilium longiflorum | unclassified | [ |
| Ls | LAS | Initiation of the axillary meristem | tomato | HAM, LAS, SCL4/7, SCR, SCL9, SCL28, DELLA, SHR, PAT1, Os4, Os19, GRAS37, and Pt20 | [ |
| PeSCL7 | SCL4/7 | Enhanced drought tolerance and salt tolerance of transgenic Arabidopsis plants | Populus euphratica | unclassified | [ |
| AtSCR | SCR | Involved in radial root morphology and growth | A. thaliana | HAM, LISCL, LAS, SCL4/7, SCR, DLT, SCL3, DELLA, SHR, and PAT1 | [ |
| AtSHR | SHR | Involved in radial root morphology and growth | A. thaliana | HAM, LISCL, LAS, SCL4/7, SCR, DLT, SCL3, DELLA, SHR, and PAT1 | [ |
| PtSHR1 | SHR | Increased growth rates | Populus tomentosa | Os19, HAM, Os4, Pt20, DLT, AtSCl3, AtSHR, AtPAT1, AtSCR, AtSCL4/7, AtLAS, DELLA, and LISCL | [ |
| DLT | DLT | Involved in brassinoltone signaling | rice | LISCL, SHR, DELLA, SCL3, PAT1, SCR, SCL4/7, LAS, Os19, HAM, Os4, and DLT | [ |
| AtSCL3 | SCL3 | Integrated multiple signals during Arabidopsis root cell elongation | A. thaliana | HAM, LISCL, LAS, SCL4/7, SCR, DLT, SCL3, DELLA, SHR, and PAT1 | [ |
| AtRGA | DELLA | Modulated jasmonate signaling via competitive binding to JAZs | A. thaliana | HAM, LISCL, LAS, SCL4/7, SCR, DLT, SCL3, DELLA, SHR, and PAT1 | [ |
| BdSLR1 BdSLRL1 | DELLA | Play a role in plant growth via the GA signal pathway | Brachypodium distachyon | HAM, PAT1, SHR, DELLA, SCL3, SCL4/7, LAS, SCR, DLT, and LISCL | [ |
| AtPAT1 | PAT1 | Involved in signaling in Arabidopsis photochromes | A. thaliana | HAM, LISCL, LAS, SCL4/7, SCR, DLT, SCL3, DELLA, SHR, and PAT1 | [ |
| GmGRAS37 | PAT1 | Improved resistance to drought and salt stresses | soybean | AtSCL4/7, Os19, Os4, HAM, DELLA, DLT, AtPAT1, LISCL, AtSCR, AtSCL3, and AtSHR | [ |
| StGRAS9 | PAT1 | Responded to plant hormones IAA, ABA, and GA3 treatment | potato | DELLA, LAS, HAM, PATI, SCR, LISCL, SHR, and SCL3 | [ |
Physicochemical properties and subcellular localization analyses of the GRAS gene family in T. cacao.
| Gene Name | Gene ID | Physicochemical Characteristics | SL | ORF | ||||
|---|---|---|---|---|---|---|---|---|
| PI | MW (Da) | Length (aa) | Instability Index | Aliphatic Index | ||||
| TcGRAS1 | TCM_000399 | 5.00 | 67,286.55 | 608 | 54.24 | 81.73 | endomembrane system | 1827 |
| TcGRAS2 | TCM_000435 | 6.80 | 77,476.49 | 684 | 50.67 | 84.71 | nucleus | 2055 |
| TcGRAS3 | TCM_000764 | 5.46 | 50,905.28 | 456 | 41.68 | 92.43 | chloroplast | 1371 |
| TcGRAS4 | TCM_000801 | 5.59 | 57,001.87 | 505 | 52.26 | 70.32 | chloroplast | 1518 |
| TcGRAS5 | TCM_002021 | 5.42 | 48,046.37 | 441 | 56.70 | 91.54 | nucleus | 1326 |
| TcGRAS6 | TCM_002319 | 5.57 | 63,858.30 | 569 | 55.85 | 79.17 | nucleus | 1710 |
| TcGRAS7 | TCM_003984 | 5.84 | 57,336.13 | 511 | 44.83 | 83.99 | nucleus | 1536 |
| TcGRAS8 | TCM_004818 | 6.15 | 63,796.67 | 565 | 31.39 | 85.91 | nucleus | 1698 |
| TcGRAS9 | TCM_005571 | 5.41 | 79,449.91 | 730 | 62.24 | 84.47 | nucleus | 2193 |
| TcGRAS10 | TCM_007806 | 5.31 | 49,737.63 | 445 | 43.06 | 88.97 | chloroplast | 1338 |
| TcGRAS11 | TCM_010708 | 6.38 | 56,277.83 | 499 | 47.40 | 92.06 | nucleus | 1500 |
| TcGRAS12 | TCM_010965 | 5.66 | 67,186.67 | 615 | 39.21 | 80.93 | nucleus | 1848 |
| TcGRAS13 | TCM_014574 | 6.04 | 57,510.78 | 519 | 52.58 | 90.02 | nucleus | 1560 |
| TcGRAS14 | TCM_015228 | 5.76 | 67,975.59 | 628 | 45.58 | 77.82 | nucleus | 1887 |
| TcGRAS15 | TCM_015991 | 5.69 | 87,113.44 | 795 | 55.54 | 80.34 | nucleus | 2388 |
| TcGRAS16 | TCM_016186 | 5.68 | 74,493.64 | 671 | 52.33 | 79.79 | nucleus | 2016 |
| TcGRAS17 | TCM_017269 | 5.67 | 72,460.97 | 654 | 53.85 | 78.35 | nucleus | 1965 |
| TcGRAS18 | TCM_017746 | 5.88 | 86,321.23 | 795 | 53.09 | 81.66 | nucleus | 2388 |
| TcGRAS19 | TCM_018964 | 6.92 | 46,686.93 | 418 | 44.25 | 94.07 | nucleus | 1257 |
| TcGRAS20 | TCM_019165 | 5.27 | 71,951.43 | 655 | 49.47 | 81.62 | nucleus | 1968 |
| TcGRAS21 | TCM_019414 | 5.43 | 39,709.90 | 347 | 49.00 | 94.96 | nucleus | 1044 |
| TcGRAS22 | TCM_019956 | 6.08 | 191,183.51 | 1659 | 47.01 | 80.49 | nucleus | 4980 |
| TcGRAS23 | TCM_019978 | 5.82 | 51,125.89 | 457 | 46.55 | 79.63 | chloroplast | 1374 |
| TcGRAS24 | TCM_021350 | 5.67 | 50,849.34 | 457 | 33.62 | 93.09 | nucleus | 1374 |
| TcGRAS25 | TCM_021351 | 5.08 | 67,410.79 | 600 | 47.40 | 83.42 | nucleus | 1803 |
| TcGRAS26 | TCM_021618 | 4.86 | 61,003.74 | 538 | 49.81 | 70.91 | nucleus | 1617 |
| TcGRAS27 | TCM_021920 | 6.17 | 85,337.10 | 755 | 52.92 | 73.09 | nucleus | 2268 |
| TcGRAS28 | TCM_029136 | 6.30 | 64,717.65 | 582 | 46.26 | 74.60 | nucleus | 1749 |
| TcGRAS29 | TCM_030393 | 5.89 | 61,290.47 | 548 | 52.83 | 79.00 | nucleus | 1647 |
| TcGRAS30 | TCM_030498 | 6.19 | 49,455.27 | 438 | 55.76 | 96.74 | nucleus | 1317 |
| TcGRAS31 | TCM_030733 | 5.56 | 60,140.03 | 540 | 58.44 | 79.35 | nucleus | 1623 |
| TcGRAS32 | TCM_031132 | 6.13 | 66,239.71 | 596 | 52.40 | 80.54 | nucleus | 1791 |
| TcGRAS33 | TCM_033446 | 4.94 | 58,582.29 | 521 | 39.07 | 86.91 | nucleus | 1566 |
| TcGRAS34 | TCM_035069 | 5.28 | 53,980.68 | 487 | 40.15 | 85.15 | nucleus | 1464 |
| TcGRAS35 | TCM_035362 | 5.06 | 63,856.24 | 570 | 49.25 | 82.14 | nucleus | 1713 |
| TcGRAS36 | TCM_036707 | 6.31 | 52,208.63 | 457 | 43.57 | 86.81 | nucleus | 1374 |
| TcGRAS37 | TCM_037975 | 5.62 | 60,414.53 | 548 | 49.30 | 87.08 | nucleus | 1647 |
| TcGRAS38 | TCM_040833 | 5.61 | 52,722.63 | 470 | 50.42 | 98.81 | nucleus | 1413 |
| TcGRAS39 | TCM_041810 | 6.33 | 79,445.42 | 705 | 46.24 | 71.65 | nucleus | 2118 |
| TcGRAS40 | TCM_041812 | 6.17 | 78,589.84 | 690 | 51.06 | 80.39 | nucleus | 2073 |
| TcGRAS41 | TCM_041813 | 5.40 | 75,260.51 | 666 | 53.55 | 81.14 | nucleus | 2001 |
| TcGRAS42 | TCM_041814 | 5.16 | 89,125.20 | 790 | 45.83 | 71.96 | nucleus | 2373 |
| TcGRAS43 | TCM_041815 | 6.30 | 92,905.70 | 829 | 46.81 | 70.95 | nucleus | 2490 |
| TcGRAS44 | TCM_042194 | 4.63 | 60,211.98 | 537 | 45.66 | 85.74 | nucleus | 1614 |
| TcGRAS45 | TCM_042392 | 5.80 | 54,285.57 | 483 | 62.33 | 96.15 | nucleus | 1452 |
| TcGRAS46 | TCM_042705 | 5.62 | 83,092.65 | 737 | 52.65 | 76.73 | nucleus | 2214 |
Tandem duplication in TcGRAS genes and corresponding Ka, Ks, and Ka/Ks values.
| Tandem Duplication | Chromosome Name | Ka | Ks | Ka/Ks |
|---|---|---|---|---|
| TcGRAS24 and TcGRAS25 | Chr04 | 5.21 | 2.66 | 1.96 |
| TcGRAS42 and TcGRAS43 | Chr09 | 5.62 | 1.41 | 3.99 |
Supplementary Materials
The following supporting information can be downloaded at:
References
1. Riano-Pachon, D.M.; Ruzicic, S.; Dreyer, I.; Mueller-Roeber, B. PlnTFDB: An integrative plant transcription factor database. BMC Bioinform.; 2007; 8, 42. [DOI: https://dx.doi.org/10.1186/1471-2105-8-42] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/17286856]
2. Liu, B.; Sun, Y.; Xue, J.; Jia, X.; Li, R. Genome-wide characterization and expression analysis of GRAS gene family in pepper (Capsicum annuum L.). PeerJ; 2018; 6, e4796. [DOI: https://dx.doi.org/10.7717/peerj.4796] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/29868257]
3. Song, Y.; Xuan, A.; Bu, C.; Ci, D.; Tian, M.; Zhang, D. Osmotic stress-responsive promoter upstream transcripts (PROMPTs) act as carriers of MYB transcription factors to induce the expression of target genes in Populus simonii. Plant Biotechnol. J.; 2019; 17, pp. 164-177. [DOI: https://dx.doi.org/10.1111/pbi.12955] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/29797449]
4. Laurenzio, L.D.; Wysocka-Diller, J.; Malamy, J.E.; Pysh, L.; Helariutta, Y.; Freshour, K.; Hahn, M.G.; Feldmann, K.A.; Benfey, P.N. The SCARECROW gene regulates an asymmetric cell division that is essential for generating the radial organization of the Arabidopsis root. Cell; 1996; 86, pp. 423-433. [DOI: https://dx.doi.org/10.1016/S0092-8674(00)80115-4]
5. Peng, J.; Carol, P.; Richards, D.E.; King, K.E.; Cowling, R.J.; Murphy, G.P.; Harberd, P.H. The Arabidopsis GAI gene defines a signaling pathway that negatively regulates gibberellin responses. Genes Dev.; 1997; 11, pp. 3194-3205. [DOI: https://dx.doi.org/10.1101/gad.11.23.3194]
6. Silverstone, A.L.; Ciampaglio, C.N.; Sun, T. The Arabidopsis RGA gene encodes a transcriptional regulator repressing the gibberellin signal transduction pathway. Plant Cell; 1998; 10, pp. 155-169. [DOI: https://dx.doi.org/10.1105/tpc.10.2.155]
7. Bolle, C. The role of GRAS proteins in plant signal transduction and development. Planta; 2004; 218, pp. 683-692. [DOI: https://dx.doi.org/10.1007/s00425-004-1203-z]
8. Tian, C.; Wan, P.; Sun, S.; Li, J.; Chen, M. Genome-wide analysis of the GRAS gene family in Rice and Arabidopsis. Plant Mol. Biol.; 2004; 54, pp. 519-532. [DOI: https://dx.doi.org/10.1023/B:PLAN.0000038256.89809.57]
9. Liu, X.; Widmer, A. Genome-wide comparative analysis of the GRAS gene family in Populus, Arabidopsis and Rice. Plant Mol. Biol. Rep.; 2014; 32, pp. 1129-1145. [DOI: https://dx.doi.org/10.1007/s11105-014-0721-5]
10. Wang, T.; Yu, T.; Fu, J.; Su, H.; Chen, J.; Zhou, Y.; Chen, M.; Guo, J.; Ma, Y.; Wei, W. et al. Genome-wide analysis of the GRAS gene family and functional identification of GmGRAS37 in drought and salt tolerance. Front. Plant Sci.; 2020; 11, 604690. [DOI: https://dx.doi.org/10.3389/fpls.2020.604690]
11. Huang, W.; Xian, Z.; Kang, X.; Tang, N.; Li, Z. Genome-wide identification, phylogeny and expression analysis of GRAS gene family in tomato. BMC Plant Biol.; 2015; 15, 209. [DOI: https://dx.doi.org/10.1186/s12870-015-0590-6] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/26302743]
12. Tang, Z.J.; Song, N.; Peng, W.Y.; Yang, Y.; Qiu, T.; Huang, C.T.; Dai, L.Y.; Wang, B. Genome identification and expression analysis of GRAS family related to development, hormone and pathogen stress in Brachypodium distachyon. Front. Sustain. Food Syst.; 2021; 5, 675177. [DOI: https://dx.doi.org/10.3389/fsufs.2021.675177]
13. Wang, S.; Zhang, N.; Zhu, X.; Yang, J.; Li, S.; Che, Y.; Liu, W.; Si, H. Identification and expression analysis of StGRAS gene family in potato (Solanum tuberosum L.). Comput. Biol. Chem.; 2019; 80, pp. 195-205. [DOI: https://dx.doi.org/10.1016/j.compbiolchem.2019.03.020] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/30978571]
14. Li, X.; Duan, X.; Jiang, H.; Sun, Y.; Tang, Y.; Yuan, Z.; Guo, J.; Liang, W.; Chen, L.; Yin, J. et al. Genome- wide analysis of basic/helix-loop-helix transcription factor family in rice and Arabidopsis. Plant Physiol.; 2006; 141, pp. 1167-1184. [DOI: https://dx.doi.org/10.1104/pp.106.080580] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/16896230]
15. Hirsch, S.; Oldroyd, G.E. GRAS-domain transcription factors that regulate plant development. Plant Signal. Behav.; 2009; 4, pp. 698-700. [DOI: https://dx.doi.org/10.4161/psb.4.8.9176] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/19820314]
16. Sun, X.; Jones, W.T.; Rikkerink, E.H. GRAS proteins: The versatile roles of intrinsically disordered proteins in plant signalling. Biochem. J.; 2012; 442, pp. 1-12. [DOI: https://dx.doi.org/10.1042/BJ20111766]
17. Stuurman, J.; Jäggi, F.; Kuhlemeier, C. Shoot meristem maintenance is controlled by a GRAS-gene mediated signal from differentiating cells. Genes Dev.; 2002; 16, pp. 2213-2218. [DOI: https://dx.doi.org/10.1101/gad.230702]
18. Morohashi, K.; Minami, M.; Takase, H.; Hotta, Y.; Hiratsuka, K. Isolation and characterization of a novel GRAS gene that regulates meiosis-associated gene expression. J. Biol. Chem.; 2003; 278, pp. 20865-20873. [DOI: https://dx.doi.org/10.1074/jbc.M301712200]
19. Schumacher, K.; Schmitt, T.; Rossberg, M.; Schmitz, G.; Theres, K. The Lateral suppressor (Ls) gene of tomato encodes a new member of the VHIID protein family. Proc. Natl. Acad. Sci. USA; 1999; 96, pp. 290-295. [DOI: https://dx.doi.org/10.1073/pnas.96.1.290]
20. Ma, H.; Liang, D.; Shuai, P.; Xia, X.; Yin, W. The salt- and drought-inducible poplar GRAS protein SCL7 confers salt and drought tolerance in Arabidopsis thaliana. J. Exp. Bot.; 2010; 61, pp. 4011-4019. [DOI: https://dx.doi.org/10.1093/jxb/erq217] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/20616154]
21. Helariutta, Y.; Fukaki, H.; Wysocka-Diller, J.; Nakajima, K.; Jung, J.; Sena, G.; Hauser, M.T.; Benfey, P.N. The SHORT-ROOT gene controls radial patterning of the Arabidopsis root through radial signaling. Cell; 2000; 101, pp. 555-567. [DOI: https://dx.doi.org/10.1016/S0092-8674(00)80865-X] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/10850497]
22. Wang, J.; Andersson-Gunneras, S.; Gaboreanu, I.; Hertzberg, M.; Tucker, M.R.; Zheng, B.; Lesniewska, J.; Mellerowicz, E.J.; Laux, T.; Sandberg, G. et al. Reduced expression of the SHORT-ROOT gene increases the rates of growth and development in hybrid poplar and Arabidopsis. PLoS ONE; 2011; 6, e28878. [DOI: https://dx.doi.org/10.1371/journal.pone.0028878] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/22194939]
23. Tong, H.; Jin, Y.; Liu, W.; Li, F.; Fang, J.; Yin, Y.; Qian, Q.; Zhu, L.; Chu, C. DWARF AND LOW-TILLERING, a new member of the GRAS family, plays positive roles in brassinosteroid signaling in rice. Plant J.; 2009; 58, pp. 803-816. [DOI: https://dx.doi.org/10.1111/j.1365-313X.2009.03825.x] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/19220793]
24. Heo, J.O.; Chang, K.S.; Kim, I.A.; Lee, M.H.; Lee, S.A.; Song, S.K.; Lee, M.M.; Lim, J. Funneling of gibberellin signaling by the GRAS transcription regulator SCARECROW-LIKE 3 in the Arabidopsis root. Proc. Natl. Acad. Sci. USA; 2011; 108, pp. 2166-2171. [DOI: https://dx.doi.org/10.1073/pnas.1012215108] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/21245304]
25. Hou, X.; Lee, L.; Xia, K.; Yan, Y.; Yu, H. DELLAs modulate jasmonate signaling via competitive binding to JAZs. Dev. Cell; 2010; 19, pp. 884-894.
26. Bolle, C.; Koncz, C.; Chua, N.H. PAT1, a new member of the GRAS family, is involved in phytochrome A signal transduction. Genes Dev.; 2000; 14, pp. 1269-1278. [DOI: https://dx.doi.org/10.1101/gad.14.10.1269]
27. Mustiga, G.M.; Gezan, S.A.; Phillips-Mora, W.; Arciniegas-Leal, A.; Mata-Quirós, A.; Motamayor, J.C. Phenotypic description of Theobroma cacao L. for yield and vigor traits from 34 hybrid families in Costa Rica based on the genetic basis of the parental population. Front. Plant Sci.; 2018; 9, 808. [DOI: https://dx.doi.org/10.3389/fpls.2018.00808]
28. Corti, R.; Flammer, A.J.; Hollenberg, N.K.; Lüscher, T.F. Cocoa and cardiovascular health. Circulation; 2009; 119, pp. 1433-1441. [DOI: https://dx.doi.org/10.1161/CIRCULATIONAHA.108.827022]
29. Argout, X.; Salse, J.; Aury, J.M.; Guiltinan, M.J.; Droc, G.; Gouzy, J.; Allegre, M.; Chaparro, C.; Legavre, T.; Maximova, S.N. The genome of Theobroma cacao. Nat. Genet.; 2011; 3, pp. 101-108. [DOI: https://dx.doi.org/10.1038/ng.736]
30. Shen, S.; Zhang, Q.; Shi, Y.; Sun, Z.; Zhang, Q.; Hou, S.; Wu, R.; Jiang, L.; Zhao, X.; Guo, Y. Genome-wide analysis of the NAC Domain transcription factor gene family in Theobroma cacao. Genes; 2019; 11, 35. [DOI: https://dx.doi.org/10.3390/genes11010035]
31. Silva Monteiro de Almeida, D.; Oliveira Jordão do Amaral, D.; Del-Bem, L.E.; Bronze Dos Santos, E.; Santana Silva, R.J.; Peres Gramacho, K.; Vincentz, M.; Micheli, F. Genome-wide identification and characterization of cacao WRKY transcription factors and analysis of their expression in response to witches’ broom disease. PLoS ONE; 2017; 12, e0187346. [DOI: https://dx.doi.org/10.1371/journal.pone.0187346] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/29084273]
32. Martins Alves, A.M.; Pereira Menezes Reis, S.; Peres Gramacho, K.; Micheli, F. The glutathione peroxidase family of Theobroma cacao: Involvement in the oxidative stress during witches’ broom disease. Int. J. Biol. Macromol.; 2020; 164, pp. 3698-3708. [DOI: https://dx.doi.org/10.1016/j.ijbiomac.2020.08.222] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/32882281]
33. Li, F.; Wu, B.; Yan, L.; Qin, X.; Lai, J. Metabolome and transcriptome profiling of Theobroma cacao provides insights into the molecular basis of pod color variation. J. Plant Res.; 2021; 134, pp. 1323-1334. [DOI: https://dx.doi.org/10.1007/s10265-021-01338-9] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/34420146]
34. Lu, J.; Wang, T.; Xu, Z.; Sun, L.; Zhang, Q. Genome-wide analysis of the GRAS gene family in Prunus mume. Mol. Gen. Genomics; 2015; 290, pp. 303-317. [DOI: https://dx.doi.org/10.1007/s00438-014-0918-1] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/25245166]
35. El-Gebali, S.; Mistry, J.; Bateman, A.; Eddy, S.R.; Luciani, A.; Potter, S.C.; Qureshi, M.; Richardson, L.J.; Salazar, G.A.; Smart, A. The Pfam protein families database in 2019. Nucleic Acids Res.; 2019; 47, pp. D427-D432. [DOI: https://dx.doi.org/10.1093/nar/gky995]
36. Larkin, M.A.; Blackshields, G.; Brown, N.P.; Chenna, R.; McGettigan, P.A.; McWilliam, H.; Valentin, F.; Wallace, I.M.; Wilm, A.; Lopez, R. Clustal W and Clustal X version 2.0. Bioinformatics; 2007; 23, pp. 2947-2948. [DOI: https://dx.doi.org/10.1093/bioinformatics/btm404]
37. Letunic, I.; Copley, R.R.; Schmidt, S.; Ciccarelli, F.D.; Doerks, T.; Schultz, J.; Ponting, C.P.; Bork, P. SMART 4.0: Towards genomic data integration. Nucleic Acids Res.; 2004; 32, pp. D142-D144. [DOI: https://dx.doi.org/10.1093/nar/gkh088]
38. Wilkins, M.R.; Gasteiger, E.; Bairoch, A.; Sanchez, J.C.; Hochstrasser, D.F. Protein identification and analysis tools in the ExPASy server. Methods Mol. Biol.; 1999; 112, pp. 531-552.
39. Kumar, S.; Stecher, G.; Tamura, K. MEGA7: Molecular evolutionary genetics analysis version 7.0 for bigger datasets. Mol. Biol. Evol.; 2016; 33, pp. 1870-1874. [DOI: https://dx.doi.org/10.1093/molbev/msw054]
40. Edgar, R.C. MUSCLE: A multiple sequence alignment method with reduced time and space complexity. BMC Bioinform.; 2004; 5, 113. [DOI: https://dx.doi.org/10.1186/1471-2105-5-113]
41. Pysh, L.D.; Wysocka-Diller, J.W.; Camilleri, C.; Bouchez, D.; Benfey, P.N. The GRAS gene family in Arabidopsis: Sequence characterization and basic expression analysis of the SCARECROW-LIKE genes. Plant J.; 1999; 18, pp. 111-119. [DOI: https://dx.doi.org/10.1046/j.1365-313X.1999.00431.x] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/10341448]
42. Kumar, S.; Stecher, G.; Li, M.; Knyaz, C.; Tamura, K. MEGA X: Molecular evolutionary genetics analysis across computing platforms. Mol. Biol. Evol.; 2018; 35, pp. 1547-1549. [DOI: https://dx.doi.org/10.1093/molbev/msy096] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/29722887]
43. Letunic, B. Interactive Tree of Life (iTOL): An online tool for phylogenetic tree display and annotation. Bioinformatics; 2007; 23, pp. 127-128. [DOI: https://dx.doi.org/10.1093/bioinformatics/btl529]
44. Bailey, T.L.; Boden, M.; Buske, F.A.; Frith, M.; Grant, C.E.; Clementi, L.; Ren, J.Y.; Li, W.F.; Noble, W.S. MEME SUITE: Tools for motif discovery and searching. Nucleic Acids Res.; 2009; 37, pp. W202-W208. [DOI: https://dx.doi.org/10.1093/nar/gkp335] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/19458158]
45. Lescot, M.; Dehais, P.; Thijs, G.; Marchal, K.; Moreau, Y.; Van de Peer, Y.; Rouze, P.; Rombauts, S. PlantCARE, a database of plant cis-acting regulatory elements and a portal to tools for in silico analysis of promoter sequences. Nucleic Acids Res.; 2002; 30, pp. 325-327. [DOI: https://dx.doi.org/10.1093/nar/30.1.325] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/11752327]
46. Wang, D.P.; Zhang, Y.B.; Zhang, Z.; Zhu, J.; Yu, J. KaKs_Calculator 2.0: A Toolkit Incorporating Gamma-Series Methods and Sliding Window Strategies. Genom. Proteom. Bioinf.; 2010; 8, pp. 77-80. [DOI: https://dx.doi.org/10.1016/S1672-0229(10)60008-3]
47. Yu, G. Using ggtree to visualize data on tree-like structures. Curr. Protoc. Bioinform.; 2020; 69, e96. [DOI: https://dx.doi.org/10.1002/cpbi.96]
48. Roy, S.W.; Penny, D. Patterns of intron loss and gain in plants: Intron loss-dominated evolution and genome-wide comparison of O. sativa and A. thaliana. Mol. Biol. Evol.; 2007; 24, pp. 171-181. [DOI: https://dx.doi.org/10.1093/molbev/msl159]
49. Rogozin, I.B.; Carmel, L.; Csuros, M.; Koonin, E.V. Origin and evolution of spliceosomal introns. Biol. Direct.; 2012; 7, 11. [DOI: https://dx.doi.org/10.1186/1745-6150-7-11]
50. Cannon, S.B.; Mitra, A.; Baumgarten, A.; Young, N.D.; May, G. The roles of segmental and tandem gene duplication in the evolution of large gene families in Arabidopsis thaliana. BMC Plant Biol.; 2004; 4, 10. [DOI: https://dx.doi.org/10.1186/1471-2229-4-10]
51. Hanada, K.; Zou, C.; Lehti-Shiu, M.D.; Shinozaki, K.; Shiu, S.H. Importance of lineage-specific expansion of plant tandem duplicates in the adaptive response to environmental stimuli. Plant Physiol.; 2008; 148, pp. 993-1003. [DOI: https://dx.doi.org/10.1104/pp.108.122457] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/18715958]
52. Jiang, S.Y.; Ma, Z.; Ramachandran, S. Evolutionary history and stress regulation of the lectin superfamily in higher plants. BMC Evol. Biol.; 2010; 10, 79. [DOI: https://dx.doi.org/10.1186/1471-2148-10-79] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/20236552]
53. Ho, C.L.; Geisler, M. Genome-wide computational identification of biologically significant cis-regulatory elements and associated transcription factors from rice. Plants; 2019; 8, 441. [DOI: https://dx.doi.org/10.3390/plants8110441] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/31652796]
© 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.