Next Article in Journal
The LIV-1 Subfamily of Zinc Transporters: From Origins to Present Day Discoveries
Next Article in Special Issue
IRAPs in Combination with Highly Informative ISSRs Confer Effective Potentials for Genetic Diversity and Fidelity Assessment in Rhododendron
Previous Article in Journal
The Effect of the Gut Microbiota on Transplanted Kidney Function
Previous Article in Special Issue
Comparative Transcriptome Analysis of CCCH Family in Roles of Flower Opening and Abiotic Stress in Osmanthus fragrans
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
IJMS
Volume 24
Issue 2
10.3390/ijms24021261
ijms-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Article

Genome-Wide Identification and Characterization of the GRF Gene Family in Melastoma dodecandrum

by
Jie Huang
,
Gui-Zhen Chen
,
Sagheer Ahmad
,
Yang Hao
,
Jin-Liao Chen
,
Yu-Zhen Zhou
,
Si-Ren Lan
,
Zhong-Jian Liu
* and
Dong-Hui Peng
*
Key Laboratory of National Forestry and Grassland Administration for Orchid Conservation and Utilization at Landscape Architecture and Arts, Fujian Agriculture and Forestry University, Fuzhou 350002, China
*
Authors to whom correspondence should be addressed.
Int. J. Mol. Sci. 2023, 24(2), 1261; https://doi.org/10.3390/ijms24021261
Submission received: 9 November 2022 / Revised: 30 December 2022 / Accepted: 6 January 2023 / Published: 9 January 2023
(This article belongs to the Special Issue Research Advances in Ornamental Plants Breeding and Biotechnology)
Browse Figures
Versions Notes

Abstract

:
Growth-regulating factor (GRF) is a kind of transcription factor unique to plants, playing an important role in the flowering regulation, growth, and development of plants. Melastoma dodecandrum is an important member of Melastomataceae, with ornamental, medicinal, and edible benefits. The identification of the GRF gene family in M. dodecandrum can help to improve their character of flavor and continuous flowering. The members of the GRF gene family were identified from the M. dodecandrum genome, and their bioinformatics, selective pressure, and expression patterns were analyzed. The results showed that there were 20 GRF genes in M. dodecandrum. Phylogenetic analysis showed that the 71 GRF genes from M. dodecandrum, Arabidopsis thaliana, Camellia sinensis, and Oryza sativa can be divided into three clades and six subclades. The 20 GRF genes of M. dodecandrum were distributed in twelve chromosomes and one contig. Furthermore, the gene structure and motif analysis showed that the intron and motif within each clade were very similar, but there were great differences among different clades. The promoter contained cis-acting elements related to hormone induction, stress, and growth and development. Different transcriptomic expression of MdGRFs indicated that MdGRFs may be involved in regulating the growth and development of M. dodecandrum. The results laid a foundation for further study on the function and molecular mechanism of the M. dodecandrum GRF gene family.

1. Introduction

Growth-regulating factors (GRFs) are transcription factors unique to plants that play a significant role in plant growth and development. They were first discovered in Oryza sativa in 2000 [1]. Recently, GRF family members have been studied in some plants, such as Zea mays [2], C. sinensis [3], Arabidopsis [4], Brassica rapa [5], Brassica napus [6], Solanu lycopersicum [7], Nicotiana tabacum [8], Manihot esculenta [9], Malus pumila [10], Cucurbitaceae [11], Brachypodium distachyon [12], Morus alba [13], Juglandaceae [14], and Nelumbo nucifera [15]. The GRF gene family plays an important role in the formation of tissues and organs in various plant biological processes, especially during the early growth period, such as leaf development [16], root growth [17], stem elongation [18], floral organ maturation [19], seed formation [20], and hormone signal transduction [21]. Previous studies have shown that the overexpression of AtGRF1 and AtGRF2 can lead to the enlargement of leaves and cotyledons, and that AtGRF8 is related to flower development in Arabidopsis [4]. The GRF gene family in S. lycopersicum is widely involved in the growth process, such as in the increased expression of SlGRF1 during fruit ripening, higher expression of SlGRF10 in biological tissue, higher expression of SlGRF4 and SlGRF8 in flower buds, and the highest expression of SlGRF6 in root [7]. The expression of AhGRF5 was higher in the rapid expansion stage of the pod in peanuts [22].
The GRF gene family has specific highly conserved domains, which are mainly located in the N-terminal, QLQ (Glu-Leu-Glu), and WRC (Trp-Arg-Cys) conserved domains [23]. The QLQ domain is highly homologous to the N-terminal of yeast SWI2/SNF2 protein and is an important protein–protein interaction domain. The interaction protein GIF (GRF-interacting factor) of AtGRF1 was identified via the yeast two-hybrid technique [24]. WRC is a kind of plant-specific motif, which is responsible for the interaction between transcription factors and DNA in the nucleus. It contains a DNA-binding nucleus localization signal region and zinc finger structure (C-X9-C-X10-C-X2-H), which regulates the expression of downstream target genes [2,4,5,8,25]. However, research on the GRF gene family in M. dodecandrum has not been reported.
M. dodecandrum is a crucial subshrub in the Melastoma genus. It is mainly distributed in southern China. It has great ornamental value and is also used as a folk herbal medicine. The plants of M. dodecandrum are low and creeping, with thick leaves close to the ground which can form a flat and dense ground cover. Their leaves, flowers, and fruits show variable colors throughout the year. It has high ornamental value and has resistance to shade and trampling. It can be used as an excellent ground cover plant in the popularization and application of landscape architecture [26]. The fruit of M. dodecandrum is a kind of juicy berry with a highly nutritive value containing vitamin C, crude protein, cyanin, and so on [27,28]. Studying the GRF family can do a great deal to understand the growth and development pattern of this species and improve its character. Therefore, we report the identification and expression analysis of the GRF gene family in M. dodecandrum.

2. Results

2.1. Identification and Physicochemical Property Analysis of GRF

Our study used the amino acid sequences of Arabidopsis and rice GRF genes as a reference. Twenty GRF genes were identified. Based on the analysis of the physicochemical properties of the GRF gene family of M. dodecandrum, it was found that the physicochemical properties of each member were different. The length of the GRF gene family of M. dodecandrum ranged from 277 to 637 aa (Table 1). The molecular weight was from 29,999.53 to 68,659.67 Da. The isoelectric point of the GRF gene family was from 5.94 to 9.3. The instability index was from 44.32 to 74.3. The grand average of hydropathicity was from −0.859 to −0.475.

2.2. Conserved Domain and Gene Structure Analysis of GRF

The analysis of the conserved domain showed that all of the GRF genes of M. dodecandrum had two completely conserved domains: WRC (Figure 1A,B) and QLQ (Figure 1C,D). Ten conserved motifs in the GRF gene family were identified using MEME online tools (Figure 2A). The results showed that all genes of the GRF family had two typical motifs, motif 1 (WRC) and motif 2 (QLQ), indicating that these two motifs were relatively conservative among the GRF gene family (Figure 1 and Figure 2A). With the exception of MdGRF4 and MdGRF20, all members of the GRF gene family had conserved motif 3. Except for MdGRF17, MdGRF12, and MdGRF11, the other 17 GRF genes contained motif 4. MdGRF10 and MdGRF7 contained the least conservative motifs, indicating that the GRF gene family of M. dodecandrum may have a complex evolutionary process.
The gene structure of the GRF gene family showed diversity (Figure 2B). The number of GRF genes’ exons of M. dodecandrum ranged from 3–5. Moreover, seven genes (MdGRF2, MdGRF3, MdGRF6, MdGRF5, MdGRF14, MdGRF17, and MdGRF7) contained two introns, MdGRF1 contained four introns, and the other GRF genes contained three introns.

2.3. Cis-acting element prediction of GRF gene family

The results of the GRF gene family of promoter acting element prediction of the upstream 2000 bp sequence showed that there were a variety of environmental and stress response elements in the GRF gene family of M. dodecandrum (Figure 3). All MdGRF genes contained light response elements (307), STREs (239), MYB biding sites (158), and MYC biding sites (116); 95% of MdGRF genes contained methyl jasmonate response element (141), abscisic acid responsiveness element (74), and gibberellin-responsive element (44); 90% of MdGRF genes contained WRE3 element (58), A-box (40), and W-box (38); 85% of MdGRF genes contained anaerobic induction element (43) and anoxic specific inducibility element (40); 80% of MdGRF genes had salicylic acid responsiveness elements (79); 75% of MdGRF genes had auxin-responsive element (35) and meristem expression element (34); 70% of MdGRFs contained DRE element (24); 65% MdGRFs had zein metabolism element (23); 55% MdGRF genes had drought inducibility element (21); 50% of MdGRFs contained low-temperature responsiveness element (21), ERE element (14), MYBHv1 binding site element (14), and endosperm expression element (10); 35% of MdGRF genes had defense and stress responsiveness element (8); 30% of MdGRF genes had circadian control element (6); 25% of MdGRF genes had ABRE3a element (7); 20% of MdGRFs had WUN-motif (4); and 10% of MdGRFs had CARE element (4). Furthermore, only 5% MdGRF genes had cell cycle regulation element (1), elicitor-mediated activation element (1), endosperm-specific negative expression element (1), differentiation of the palisade mesophyll element (1), and JERE element (1).

2.4. Phylogeny and Evolution Analysis of GRF Gene Family

The phylogenetic tree was constructed with GRF members of M. dodecandrum (20), C. sinensis (15), A. thaliana (9), E. grandis (8), P. granatum (8), and O. sativa (9) [1]. The result showed that these 71 GRF genes could be divided into three clades (Clades I-III) and six subclades (subclades a–f) (Figure 4). Clade I contained two subclades (a–b), Clade II only had one subclade (c), and Clade II contained three subclades (d–f). In subclade a, there were four MdGRFs, two EgGRFs, two AtGRFs, two OsGRFs, two CsGRFs, and one PgGRF. There were three MdGRFs, one EgGRF, three OsGRFs, three CsGRFs, one PgGRF, and no AtGRF found in subclade b. In subclade c, there were seven MdGRFs, three EgGRFs, three OsGRFs, two CsGRFs, and two PgGRFs, while there was no OsGRF in this subclade. Subclade d had three MdGRFs, one EgGRF, two CsGRFs, one PgGRF, and two AtGRFs, while there was no OsGRF in this subclade. Subclade e contained three OsGRFs, one AtGRF, one PgGRF, and one CsGRF, but no MdGRF or EgGRF were found in this subclade. There were three genes of MdGRFs, one EgGRF, five CsGRFs, four OsGRFs, three CsGRFs, two PgGRFs, and two AtGRFs in the subclade f.

2.5. Gene Chromosome Mapping and Collinearity Analysis

Chromosome mapping showed that 20 GRF genes were unevenly distributed on 12 chromosomes of M. dodecandrum (Figure 5). There were four genes (MdGRF1, MdGRF2, MdGRF3, and MdGRF4) in Chromosome 1. Chromosome 2, Chromosome 3, Chromosome 9, and Chromosome 12 all contained two genes. Only one gene was present on Chromosome 4, Chromosome 5, Chromosome 6, Chromosome 7, Chromosome 8, Chromosome 10, and Chromosome 11. The remaining one gene was located in the contig 164.
In order to understand the evolutionary mechanism of the GRF gene family of M. dodecandrum, we analyzed the collinear relationship between M. dodecandrum and E. grandis, and P. granatum (Figure 6). The results show that 22 pairs of collinearity genes of GRF were between M. dodecandrum and P. granatum, followed by M. dodecandrum and E. grandis (19 pairs), and the least was M. dodecandrum and A. thaliana (15 pairs).
According to the result of evolution analysis, the values of Ka, Ks, and Ka/Ks were obtained (Table 2). Nineteen gene pairs were identified using Tbtools. The value of Ka/Ks of each pair ranged from 0.155721552 to 0.44840491 (Ka/Ks < 1). This result indicated that all of them had undergone strong purifying selection.

2.6. Expression Pattern Analysis of GRF Gene Family

Based on the transcriptome data of M. dodecandrum, the tissue expression pattern of the GRF gene family was analyzed, and nine samples including root, stem, leaf, flower bud, medium flower bud, mature flower, small fruit, medium fruit, and big fruit were selected for prediction. The result showed that the MdGRF genes were expressed differently in the nine different samples. The GRF genes were expressed in root, stem, leaf, flower bud, medium flower bud, mature flower, small fruit, medium fruit, and big fruit, especially in the stem, leaf, flower bud, medium flower bud, and small fruit (Figure 7). MdGRF19 was highly expressed in the stem and leaf, and showed normal expression in the flower bud and small fruit and low expression in the other samples. MdGRF17 was highly expressed in the root, while it showed a medium or low expression in the other samples. MdGRF8 showed significant expression in the leaf and big fruit, while medium or low expression was observed in the other samples. A high expression of MdGRF18 was observed in the stem, flower bud, and medium flower bud, while a medium or low expression was seen in the other samples. Nearly the same pattern was observed for MdGRF1, MdGRF2, MdGRF3, MdGRF5, MdGRF7, and MdGRF11, which had high expression in flower bud and stem, and were nearly not expressed in root, leaf, medium flower bud, mature flower, small fruit, medium fruit, and big fruit. However, the expression of MdGRF6 was very low in the nine different samples. It is speculated that the difference in its expression may be related to the mechanism of plant growth and the development being regulated by the GRF gene family.
RT-qPCR was performed to assess the accuracy of transcriptome sequencing of all nine samples in MdGRF1, MdGRF2, MdGRF3, MdGRF7, MdGRF15, and MdGRF19. The result of RT-qPCR showed high expression in the flower bud, stem, and small fruit (Figure 8). Three genes (MdGRF1, MdGRF2, and MdGRF3) showed nearly the same between the result of RT-qPCR and transcriptome data, and MdGRF7 and MdGRF15 were nearly the same. However, with the gene of MdGRF19, it was expressed the highest in the stem according to the transcriptome data, and was medium expressed in the flower. However, according to the result of RT-qPCR, MdGRF19 was expressed the highest in the flower bud. These differences may be caused by the imperfect correlation between sequencing and RT-qPCR samples.

2.7. Subcellular Localization Analysis of MdGRF19

In this study, all MdGRF proteins were predicted to target the nucleus (Table 1). To identify the subcellular localization of MdGRF proteins, we random cloned the MdGRF19. One fusion vector was constructed and then transformed into tobacco leaf. The results show that 35S::MdGRF19-GFP was detected as being localized to the nuclear and membrane (Figure 9), which was consistent with the prediction result (Table 1). Moreover, the GFP of the empty protein (35S::GFP) was used as a control group, which was localized to nuclear and cell membranes.

3. Discussion

Plant GRF transcription factors play an important regulatory role in plant growth [29]. Our study identified 20 GRF genes of M. dodecandrum. Remarkably, the number of GRF genes of M. dodecandrum was far more than A. thaliana (9), citrus (9), E. grandis (8), P. granatum (8), and O. sativa (9) [1,24], revealing that the GRF genes of M. dodecandrum might have undergone large-scale duplication events during evolution. An analysis of the physicochemical properties of GRF in M. dodecandrum show that their GRAVY was less than zero, indicating that the MdGRFs belong to hydrophilic protein. Moreover, the instability index of MdGRFs was more than 40, showing that the structures of MdGRF proteins were unstable.
The previous study indicated that the structure of exon-intron plays an important role in understanding the relationships between evolutionary and functional differentiation [30,31,32]. Furthermore, exon or intron gain/loss events create the gene structure divergence and functional differentiation [33,34]. The structural analysis of MdGRF genes showed that there was little difference in the number and distribution of exons (3–5) and introns (2–4), which indicated that the GRF gene family of M. dodecandrum was highly conserved in the process of evolution. There are two to three introns in A. thaliana [4]; one to three introns are found in Brachypodium distachyon [12], while two to four introns are found in O. sativa [1], soybean [35], and wheat [36], which is consistent with M. dodecandrum. Moreover, different plants contain different numbers of exons; for example, AtGRF07 contains five exons, while MdGRF2 only has two exons. The results showed the diversity in the GRF genes of different plants. Two conserved domains (QLQ/WRC) at the N-terminal were predicted in the MdGRF protein sequences, which was consistent with the results of the study that found that QLQ plays a role in the protein–protein interaction and WRC effectively regulates the transcription of DNA binding c. These results indicated that MdGRF proteins were evolutionarily conserved in plants.
The result of phylogenic analysis showed that 71 GRF genes could be divided into three clades and six subclades. However, the GRF gene family of M. dodecandrum and A. thaliana was only divided into five subclades, and no gene was located in the subclades e and b, respectively. Moreover, OsGRF only had four subclades, and lost subclades c and d. We speculated that this phenomenon may be caused by a special gene expansion event (lost or obtained) during the evolutionary process [3]. The phylogenic analysis of GRF gene families in M. dodecandrum, A. thaliana, C. sinensis, E. grandis, P. granatum, and O. sativa showed that M. dodecandrum had higher homology with P. granatum and E. grandis, but a distant genetic relationship with O. sativa and C. sinensis, which revealed that some ancestor GRF genes existed before the divergence of M. dodecandrum, P. granatum, and E. grandis during evolution. Additionally, these differences also indicated that there may be structural and functional differentiation of the GRF family in dicot and monocot plants, which needs further study. Strong synteny was detected in the M. dodecandrum and P. granatum, followed by M. dodecandrum and E. grandis, and the last was the M. dodecandrum and Arabidopsis genomes. The Ka/Ks of the 19 gene pairs indicated that strong purifying selection may be largely responsible for maintaining the functions of GRF proteins, the same with A. thaliana and O. sativa [1,24].
GRFs are a key regulator of plant growth and development [37,38]. Previous study has shown that GRFs endow proliferation and meristem potential in the process of organogenesis [39]. Study has also shown that the GRF gene can regulate leaf area by controlling cell proliferation, thus enhancing the adaptation to stress conditions such as drought and high temperature [40]. The cis-acting element analysis showed that MdGRFs members could be transcribed and expressed under different stresses such as low temperature, drought, and anaerobic stress, and regulated under gibberellin and auxin stimulation signals. Some genes were involved in the growth regulation of meristem, embryo, and seed growth. The previous studies show that the expression level of the GRF gene family in growing zones is significantly higher than that in mature tissues [3,4,24], such as the OsGRFs in rice being strongly expressed in buds, immature leaves, and flower buds [25]. The expression of the GRF gene family is highest in the tender organ of Camellia sinensis, followed by the stems and immature leaves, and is hardly expressed in the root and mature flower [3]. In M. dodecandrum, the expression analysis according to RT-qPCR and transcriptome data of the GRF gene family showed that most MdGRFs were highly expressed in stem, flower bud, and small fruit, while MdGRFs were nearly not expressed in mature flower, root, and mature fruit, which is similar to previous studies on other species [3,25]. The high expression in the tender organ of the GRF gene family in M. dodecandrum indicated that MdGRFs may play an important role in regulating the development of plants.

4. Materials and Methods

4.1. Data Sources

Tender leaf, tender root, tender stem, small fruit, medium fruit, big fruit, flower bud, medium flower bud, and mature flower of wild M. dodecandrum were sampled for qPCR. The genome, transcriptome data, and GFF file used in this study follow Hao et al. [41]. Hisat [42] and Stringtie2 [43] were used to align and assemble the transcriptome data of M. dodecandrum.

4.2. Identification and Physicochemical Property Analysis of GRF Gene Family in M. dodecandrum

A. thaliana, E. grandis, P. granatum, and O. sativa GRF gene families were used as reference. GRF proteins of A. thaliana and O. sativa were downloaded from NCBI (https://www.ncbi.nlm.nih.gov/orffinder/, accessed on 17 May 2022). The Blastp alignment of GRF gene family of M. dodecandrum was carried out using TBtools [44], and the E value was less than le-5. The obtained MdGRF protein sequences were listed on CDD (https://www.ncbi.nlm.nih.gov/cdd, accessed on 8 May 2022) to detect their domains, and the protein sequences without QLQ or WRC domain were removed. In addition, two pfam seed models WRC (PF08879) and QLQ (PF08880) were obtained from the online database (http://pfam.xfam.org/, accessed on 8 May 2022) and were used for building a hidden Markov model (HMM) file using HMMER3 software with default parameters. The HMM search program was performed to search for GRF genes from hmm file generated in the previous step. We compared the results of HMM and BLASTP, and removed the repeat genes.
The MdGRF protein sequences were run on the ExPasy website (http://au.expasy.org/tool.html, accessed on 5 May 2022) and analyzed using the Compute pI/MW tool, and a variety of physicochemical properties of GRF protein were obtained, including amino acid length, molecular weight, isoelectric point, and so on [45]. Then, the CELLO v2.5 software (http://cello.life.nctu.edu.tw/, accessed on 17 May 2022) was used to predict the subcellular localization of MdGRF gene family.

4.3. Conserved Domain and Gene Structure Analysis of GRF

The gene structure of GRF family of M. dodecandrum was analyzed, and the online NCBI Conserve Domain (https://www.ncbi.nlm.nih.gov/cdd/, accessed on 17 May 2022) and MEME (https://meme-suite.org/meme/, accessed on 17 May 2022) were used to predict the conserved domain and motif of GRF gene family [46]. The results were visualized using TBtools (version 1.100, China).

4.4. Phylogeny Analysis of GRF Gene Family

Mega7 (version 7.0, America) was used to align the protein sequences of GRF genes and the phylogenetic tree (maximum likelihood) of six species (M. dodecandrum, C. sinensis, A. thaliana, E. grandis, P. granatum, and O. sativa) under a GTRGAMMA substitution model with 1000 bootstraps was constructed [47]. FigTree v1.4.3 was used to edit the tree. Lastly, adobe illustrator was used to beautify the tree.

4.5. Chromosome Mapping, Collinearity Analysis

According to the gene location information of GFF annotation file of M. dodecandrum genome, the location of GRF genes was mapped using TBtools. We also used TBtools to analyze the collinear relationship between M. dodecandrum and A. thaliana, E. grandis, and P. granatum. During evolution, genes may face various selection pressures, positive selection, neutral selection, and purifying selection, for example. This is important for studying the evolution of genes to understand the selection pressure of genes [48]. To understand the evolution of MdGRFs, Ka, Ks, and Ka/Ks were calculated. The simple Ka/Ks calculator in Tbtools (version 1.100, China) was used to analyze it.

4.6. Cis-Acting Element Prediction of GRF Gene Family

Tbtools (version 1.100, China) was used to extract the 2000 bp upstream sequences of GRF of M. dodecandrum from genomic data. The sequences were uploaded to the online website PlantCART (http://bioinformatics.psb.ugent.be/webtools/plantcare/html/, accessed on 3 May 2022) to predict the Cis-acting elements of GRF of M. dodecandrum [49]. The results of PlantCART were used to draw a picture of cis-acting element prediction.

4.7. Transcriptome Data and RT-qPCR Analysis of GRF Gene Family

In order to research the possible effect of GRF genes in different organs of M. dodecandrum, we analyzed the expression patterns of 20 GRF genes in the leaf, root, stem, small fruit, medium fruit, big fruit, flower bud, medium flower bud, and mature flower. From the transcriptome analysis, we calculated the FPKM of all nine samples, and used TBtools (version 1.100, China) to draw a heatmap.
Six genes (MdGRF1, MdGRF2, MdGRF3, MdGRF7, MdGRF15, and MdGRF19) in four subfamilies were used for RT-qPCR. Total RNA of all nine samples were extracted using the TIANGEN DP441 Reagent (TIANGEN, Beijing, China). Roche detection system (Roche, Switzerland) with SYBR green assays was used for RT-qPCR analysis. Primers and reference gene information for RT-qPCR are listed in Table S1. All experiments were performed in three biological repeats. The relative expression of MdGRFs were calculated using the 2−ΔΔCT method.

4.8. Subcellular Localization Analysis

The MdGRF19 was used further to the subcellular localization analysis. The MdGRF19 coding sequence (CDS) without stop codon was cloned into pMDC202 vector, which contained a 35s-driven green fluorescent protein (GFP) promoter. The In-Fusion cloning kit named ClonExpress® Ultra One Step Cloning Kit, which is produced by Vazyme, was used to clone. KpnI and XbaI were the choices utilized as the restriction sites of pMDC202 vector. Then, we transformed the 35S::MdGRF19-GFP into tobacco leaf, and the vector without the gene was used as a control. After 8 h of dark culture, the transformed tobacco was cultured normally (dark for eight hours at 22 °C, light for sixteen hours at 24 °C). After 48 h, the LSM710 confocal laser scanning microscope (CarlZeiss, Jena, Germany) was used to observe the GFP fluorescence signals. The primers of MdGRF19 used in this study were as follows: forwards: ttggagaggacctcgactctagaATGAGCAGCAGTGGGATGAGCAGAT; reverse: tttttctaccggtaccGATATAATGGAAAAATGAGAAAC.

5. Conclusions

This study identified the GRF gene family of M. dodecandrum and comparative analysis was performed with A. thaliana, E. grandis, and P. granatum. We studied physicochemical properties, conserved domains, gene structures, phylogeny, cis-acting element prediction chromosome mapping, and collinearity analysis of GRF of M. dodecandrum. Then, according to the transcriptome data, we speculated that the GRF gene family of M. dodecandrum may play an important role in the development and growth of M. dodecandrum, especially in the tender organ. The result of RT-qPCR also supports it. The findings of this study thus provide potential research directions to reveal the role of GRF TFs in the regulation of important ornamental traits of plants.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/ijms24021261/s1.

Author Contributions

Y.-Z.Z. and J.-L.C. designed the experiment; Z.-J.L., D.-H.P., S.-R.L. and S.A. revised the manuscript; G.-Z.C. and Y.H. performed the experiments; J.H. wrote the paper. All authors have read and agreed to the published version of the manuscript.

Funding

This research was supported by the following fundings: one is the Forestry Peak Discipline Construction Project of Fujian Agriculture and Forestry University (No. 72202200205), the other is the Innovation and Application Engineering Technology Research Center of Ornamental Plant Germplasm Resources in Fujian Province (No. 115-PTJH16005).

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

Not applicable.

Conflicts of Interest

The authors declare that there were no conflict of interest.

References

  1. Van der Knaap, E.; Kim, J.H.; Kende, H. A novel gibberellin-in-duced gene from rice and its potential regulatory role in stem growth. Plant Physiol. 2000, 122, 695–704. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  2. Zhang, D.-F.; Li, B.; Jia, G.-Q.; Zhang, T.-F.; Dai, J.-R.; Li, J.-S.; Wang, S.-C. Isolation and characterization of genes encoding GRF transcription factors and GIF transcriptional coactivators in maize (Zea mays L.). Plant Sci. 2008, 175, 809–817. [Google Scholar] [CrossRef]
  3. Wang, P.; Zheng, Y.; Lin, Y.; Zhou, Z.; Yang, J.; Ye, N. Genome-wide identification and expression analysis of GRF gene family in Camellia sinensis. Acta Bot. Boreal. Occident. Sin. 2019, 39, 0413–0421. [Google Scholar]
  4. Kim, J.H.; Choi, D.; Kende, H. The AtGRF family of putative transcription factors is involved in leaf and cotyledon growth in Arabidopsis. Plant J. 2003, 36, 94–104. [Google Scholar] [CrossRef] [PubMed]
  5. Wang, F.; Qiu, N.; Ding, Q.; Li, J.; Zhang, Y.; Li, H.; Gao, J. Genome-wide identification and analysis of the growth-regulating factor family in chinese cabbage (Brassica Rapa L. Ssp. Pekinensis). BMC Genom. 2014, 15, 807. [Google Scholar] [CrossRef] [Green Version]
  6. Ma, J.-Q.; Jian, H.-J.; Yang, B.; Lu, K.; Zhang, A.-X.; Liu, P.; Li, J.-N. Genome-wide analysis and expression profiling of the GRF gene family in oilseed rape (Brassica Napus L.). Gene 2017, 620, 36–45. [Google Scholar] [CrossRef]
  7. Khatun, K.; Robin, A.H.K.; Park, J.-I.; Nath, U.K.; Kil Kim, C.; Lim, K.-B.; Nou, I.S.; Chung, M.-Y. Molecular characterization and expression profiling of tomato GRF transcription factor family genes in response to abiotic stresses and phytohormones. Int. J. Mol. Sci. 2017, 18, 1056. [Google Scholar] [CrossRef] [Green Version]
  8. Zhang, J.; Li, Z.; Jin, J.; Xie, X.; Zhang, H.; Chen, Q.; Luo, Z.; Yang, J. Genome-wide identification and analysis of the growth-regulating factor family in tobacco (Nicotiana Tabacum). Gene 2017, 639, 117–127. [Google Scholar] [CrossRef]
  9. Shang, S.; Wu, C.; Huang, C.; Tie, W.; Yan, Y.; Ding, Z.; Xia, Z.; Wang, W.; Peng, M.; Tian, L.; et al. Genome-wide analysis of the GRF family reveals their involvement in abiotic stress response in Cassava. Genes 2018, 9, 110. [Google Scholar] [CrossRef] [Green Version]
  10. Zheng, L.; Ma, J.; Song, C.; Zhang, L.; Gao, C.; Zhang, D.; An, N.; Mao, J.; Han, M. Genome-wide identification and expression analysis of GRF genes regulating apple tree architecture. Tree Genet. Genomes 2018, 14, 54. [Google Scholar] [CrossRef]
  11. Baloglu, M.C. Genome-wide in silico identification and comparison of growth regulating factor (GRF) genes in Cucurbitaceae family. Plant Omics. 2014, 7, 260–270. [Google Scholar] [CrossRef]
  12. Filiz, E.; Koç, İ.; TombuloğLu, H. Genome-wide identification and analysis of growth regulating factor genes in Brachypodium distachyon: In silico approaches. Turkish J. Biol. 2014, 38, 296–306. [Google Scholar] [CrossRef]
  13. Rukmangada, M.S.; Sumathy, R.; Sivaprasad, V.; Naik, V.G. Genome-wide identification and characterization of growth-regulating factors in mulberry (Morus spp.). Trees 2018, 32, 1695–1705. [Google Scholar] [CrossRef]
  14. Zhang, Z.; Quan, S.; Niu, J.; Guo, C.; Kang, C.; Liu, J.; Yuan, X. Comprehensive Identification and Analyses of the GRF Gene Family in the Whole-Genome of Four Juglandaceae Species. Int. J. Mol. Sci. 2022, 23, 12663. [Google Scholar] [CrossRef] [PubMed]
  15. Chen, G.-Z.; Huang, J.; Zhou, X.-Q.; Hao, Y.; Chen, J.-L.; Zhou, Y.-Z.; Ahmad, S.; Lan, S.; Liu, Z.-J.; Peng, D.-H. Comprehensive Analysis for GRF Transcription Factors in sacred lotus (Nelumbo nucifera). Int. J. Mol. Sci. 2022, 23, 6673. [Google Scholar] [CrossRef]
  16. Horiguchi, G.; Kim, G.T.; Tsukaya, H. The transcription factor AtGRF5 and the transcription coactivator AN3 regulate cell proliferation in leaf primordia of Arabidopsis thaliana. Plant J. 2005, 43, 68–78. [Google Scholar] [CrossRef] [PubMed]
  17. Rodriguez, R.E.; Ercoli, M.F.; Debernardi, J.M.; Breakfield, N.W.; Mecchia, M.A.; Sabatini, M.; Cools, T.; De Veylder, L.; Benfey, P.N.; Palatnik, J.F. MicroRNA miR396 regulates the switch between stem cells and transit-amplifying cells in Arabidopsis roots. Plant Cell 2015, 27, 3354–3366. [Google Scholar] [CrossRef]
  18. Kemal, A.M.; Muavviz, A.; Hüseyin, U.; Emre, S.; Örenay, B.S.; Çiğdem, Y. Order-wide in silico comparative analysis and identification of growth-regulating factor proteins in Malpighiales. Turk. J. Biol. 2016, 40, 26–42. [Google Scholar] [CrossRef]
  19. Lee, S.-J.; Lee, B.H.; Jung, J.-H.; Park, S.K.; Song, J.T.; Kim, J.H. Growth-regulating factor and GRF-in-teracting factor specify meristematic cells of gynoecia and anthers. Plant Physiol. 2018, 176, 717–729. [Google Scholar] [CrossRef] [Green Version]
  20. Li, S.; Gao, F.; Xie, K.; Zeng, X.; Cao, Y.; Zeng, J.; He, Z.; Ren, Y.; Li, W.; Deng, Q.; et al. The OsmiR396c-OsGRF4-OsGIF1 regulatory module determines grain size and yield in rice. Plant Biotechnol. J. 2016, 14, 2134–2146. [Google Scholar] [CrossRef] [Green Version]
  21. Che, R.H.; Tong, H.N.; Shi, B.H.; Liu, Y.Q.; Fang, S.R.; Liu, D.P.; Xiao, Y.H.; Hu, B.; Liu, L.C.; Wang, H.R.; et al. Control of grain size and rice yield by GL2-mediated brassinosteroid responses. Nat. Plants 2005, 2, 15195. [Google Scholar] [CrossRef] [PubMed]
  22. Zhao, K.; Li, K.; Ning, L.; He, J.; Ma, X.; Li, Z.; Zhang, X.; Yin, D. Genome-wide analysis of the growth-regulating factor family in peanut (Arachis hypogaea L.). Int. J. Mol. Sci. 2019, 2, 4120. [Google Scholar] [CrossRef] [Green Version]
  23. Rodriguez, R.E.; Ercoli, M.F.; Debernardi, J.M.; Palatnik, J.F. Growth-regulating factors, a transcription factor family regulating more than just plant growth. In Plant Transcription Factors; Gonzalez, D.H., Ed.; Academic Press: Boston, MA, USA, 2016; pp. 269–280. [Google Scholar] [CrossRef]
  24. Kim, J.H.; Kende, H. A transcriptional coactivator, AtGIF1, is involved in regulating leaf growth and morphology in Arabidopsis. Proc. Natl. Acad. Sci. USA 2004, 101, 13374–13379. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  25. Choi, D.; Kim, J.H.; Kende, H. Whole genome analysis of the OsGRF gene family encoding plant-specific putative transcription activators in rice (Oryza Sativa L.). Plant Cell Physiol. 2004, 45, 897–904. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  26. He, G.F.; Xia, Y.P. A fine wild ground cover plant for flower viewing Melastoma dodecandrum. North. Hor. 2005, 3, 39. [Google Scholar]
  27. Yu, Z.C.; Lin, X.X.; Su, J.Q.; Lin, Q.J.; Chen, Z.D. Research progress of Melastoma dodecandrum. J. Anhui Agric. Sci. 2011, 39, 16557–16559. [Google Scholar]
  28. Wang, J.F.; Zhang, P.H.; Wang, Y.T.; Wang, L.H.; Li, Q.; Lin, R.C.; Zhang, J.J. Research progress on Melastoma dodecandrum. Chin. Tradit. Herb. Drugs 2018, 49, 1211–1219. [Google Scholar] [CrossRef]
  29. Tian, N.; Liu, F.; Wu, J.W.; Liu, J.P.; Li, D.; Fu, S.; Huang, Y.J.; Cheng, C.Z. Genome-wide identification and expression analysis of GRF gene family in banana. J. Fruit Sci. 2020, 2, 1821–1835. [Google Scholar] [CrossRef]
  30. Cao, D.; Wang, J.; Ju, Z.; Liu, Q.; Li, S.; Tian, H.; Fu, D.; Zhu, H.; Luo, Y.; Zhu, B. Regulations on growth and development in tomato cotyledon, flower and fruit via destruction of miR396 with short tandem target mimic. Plant Sci. 2016, 247, 1–12. [Google Scholar] [CrossRef]
  31. Liu, X.; Guo, L.-X.; Jin, L.-F.; Liu, Y.-Z.; Liu, T.; Fan, Y.-H.; Peng, S.-A. Identification and transcript profiles of citrus growth-regulating factor genes involved in the regulation of leaf and fruit development. Mol. Biol. Rep. 2016, 43, 1059–1067. [Google Scholar] [CrossRef]
  32. Büyük, I.; Aras, S. Genome-wide in silico identification, characterization and transcriptional analysis of the family of growth-regulating factors in common bean (Phaseolus vulgaris L.) subjected to polyethylene glycol-induced drought stress. Arch. Biol. Sci. 2017, 69, 5–14. [Google Scholar] [CrossRef]
  33. Xu, G.; Guo, C.; Shan, H.; Kong, H. Divergence of duplicate genes in exon–intron structure. Proc. Natl. Acad. Sci. USA 2012, 109, 1187–1192. [Google Scholar] [CrossRef] [PubMed]
  34. Hu, L.F.; Liu, S.Q. Genome-wide identification and phylogenetic analysis of the GRF gene family in cucumbers. Genet Mol. Biol. 2011, 34, 624–633. [Google Scholar] [CrossRef] [Green Version]
  35. Chen, F.; Yang, Y.; Luo, X.; Zhou, W.; Dai, Y.; Zheng, C.; Liu, W.; Yang, W.; Shu, K. Genome-wide identification of GRF transcription factors in soybean and expression analysis of GmGRF family under shade stress. BMC Plant Biol. 2019, 19, 269. [Google Scholar] [CrossRef] [PubMed]
  36. Zan, T.; Zhang, L.; Xie, T.T.; Li, L.Q. Genome-wide identification and analysis of the growth-regulating factor (GRF) gene family and GRF-interacting factor family in Triticum aestivum L. Biochem. Genet. 2020, 58, 705–724. [Google Scholar] [CrossRef]
  37. Kim, J.H.; Tsukaya, H. Regulation of plant growth and development by the growth-regulating factor and GRF-interacting factor duo. J. Exp. Bot. 2015, 66, 6093–6107. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  38. Liebsch, D.; Palatnik, J.F. MicroRNA miR396, GRF transcription factors and GIF co-regulators: A conserved plant growth regulatory module with potential for breeding and biotechnology. Curr. Opin. Plant Biol. 2020, 53, 31–42. [Google Scholar] [CrossRef]
  39. Kim, J.H. Biological roles and an evolutionary sketch of the GRF-GIF transcriptional complex in plants. BMB Rep. 2019, 52, 227–238. [Google Scholar] [CrossRef] [Green Version]
  40. Omidbakhshfard, M.A.; Proost, S.; Fujikura, U.; Mueller-Roe-ber, B. Growth-regulating factors (GRFs): A small transcription factor family with important functions in plant biology. Mol. Plant 2015, 8, 998–1010. [Google Scholar] [CrossRef] [Green Version]
  41. Hao, Y.; Zhou, Y.-Z.; Chen, B.; Chen, G.-Z.; Wen, Z.-Y.; Zhang, D.; Sun, W.-H.; Liu, D.-K.; Huang, J.; Chen, J.-L.; et al. The Melastoma dodecandrum genome and the evolution of Myrtales. J. Genet. Genom. 2022, 49, 120–131. [Google Scholar] [CrossRef]
  42. Kim, D.; Langmead, B.; Salzberg, S. Hisat: A fast spliced aligner with low memory requirements. Nat. Methods 2015, 12, 357–360. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  43. Kovaka, S.; Zimin, A.V.; Pertea, G.M.; Razaghi, R.; Salzberg, S.L.; Pertea, M. Transcriptome assembly from long-read RNA-seq alignments with StringTie2. Genome Biol. 2019, 20, 278. [Google Scholar] [CrossRef]
  44. Chen, C.J.; Chen, H.; Zhang, Y.; Thomas, H.R.; Frank, M.H.; He, Y.H.; Xia, R. TBtools: An integrative toolkit developed for interactive analyses of big biological data. Mol. Plant. 2020, 13, 1194–1202. [Google Scholar] [CrossRef] [PubMed]
  45. Artimo, P.; Jonnalagedda, M.; Arnold, K.; Baratin, D.; Csardi, G.; de Castro, E.; Duvaud, S.; Flegel, V.; Fortier, A.; Gasteiger, E.; et al. ExPASy: SIB bioinformatics resource portal. Nucleic Acids Res. 2012, 40, 597–603. [Google Scholar] [CrossRef] [PubMed]
  46. Bailey, T.L.; Boden, M.; Buske, F.A.; Frith, M.; Grant, C.E.; Clementi, L.; Ren, J.; Li, W.W.; Noble, W.S. MEME SUITE: Tools for motif discovery and searching. Nucleic Acids Res. 2009, 37, 202–208. [Google Scholar] [CrossRef]
  47. Kumar, S.; Stecher, G.; Tamura, K. Mega7: Molecular evolutionary genetics analysis version 7.0 for bigger datasets. Mol. Biol. Evol. 2016, 33, 1870–1874. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  48. Khan, N.; Fatima, F.; Haider, M.S.; Shazadee, H.; Liu, Z.; Zheng, T.; Fang, J. Genome-wide identification and expression profiling of the polygalacturonase (PG) and pectin methylesterase (PME) genes in grapevine (Vitis vinifera L.). Int. J. Mol. Sci. 2019, 20, 3180. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  49. Lescot, M.; Déhais, P.; Thijs, G.; Marchal, K.; Moreau, Y.; Van de Peer, Y.; Rouzé, 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, 325–327. [Google Scholar] [CrossRef]
Ijms 24 01261 g001 550
Figure 1. Conserved domains and Seqlogos of GRF gene family in M. dodecandrum. (A), Seqlogo of WRC (motif 1); (B), conserved sequence alignment of WRC; (C), Seqlogo of QLQ (motif 2); (D), conserved sequence alignment of QLQ.
Figure 1. Conserved domains and Seqlogos of GRF gene family in M. dodecandrum. (A), Seqlogo of WRC (motif 1); (B), conserved sequence alignment of WRC; (C), Seqlogo of QLQ (motif 2); (D), conserved sequence alignment of QLQ.
Ijms 24 01261 g001
Ijms 24 01261 g002 550
Figure 2. Motifs and gene structure of GRF gene family in M. dodecandrum. (A), motifs of MdGRF; (B), exon and intron of MdGRF.
Figure 2. Motifs and gene structure of GRF gene family in M. dodecandrum. (A), motifs of MdGRF; (B), exon and intron of MdGRF.
Ijms 24 01261 g002
Ijms 24 01261 g003 550
Figure 3. Types and numbers of cis-acting elements in promoters of M. dodecandrum GRF genes. Note: The darker the color, the greater the value.
Figure 3. Types and numbers of cis-acting elements in promoters of M. dodecandrum GRF genes. Note: The darker the color, the greater the value.
Ijms 24 01261 g003
Ijms 24 01261 g004 550
Figure 4. Phylogeny analysis among M. dodecandrum, C. sinensis, A. thaliana, E. grandis, P. granatum, and O. sativa. Note: gene name initial of ‘At’ are genes of A. thaliana, ‘Md’ are genes of M. dodecandrum, ‘Os’ are genes of O. sativa, ‘Cs’ are genes of C. sinensis, ‘Eg’ are genes of E. grandis, and ‘Pg’ are genes of P. granatum.
Figure 4. Phylogeny analysis among M. dodecandrum, C. sinensis, A. thaliana, E. grandis, P. granatum, and O. sativa. Note: gene name initial of ‘At’ are genes of A. thaliana, ‘Md’ are genes of M. dodecandrum, ‘Os’ are genes of O. sativa, ‘Cs’ are genes of C. sinensis, ‘Eg’ are genes of E. grandis, and ‘Pg’ are genes of P. granatum.
Ijms 24 01261 g004
Ijms 24 01261 g005 550
Figure 5. Chromosome localization of GRF gene family of M. dodecandrum.
Figure 5. Chromosome localization of GRF gene family of M. dodecandrum.
Ijms 24 01261 g005
Ijms 24 01261 g006 550
Figure 6. Synteny analysis of GRF gene family between M. dodecandrum and A. thaliana, E. grandis and P. granatum. Note: gray lines represent colinear gene pairs between M. dodecandrum and other species, and red lines represent colinear GRF gene pairs.
Figure 6. Synteny analysis of GRF gene family between M. dodecandrum and A. thaliana, E. grandis and P. granatum. Note: gray lines represent colinear gene pairs between M. dodecandrum and other species, and red lines represent colinear GRF gene pairs.
Ijms 24 01261 g006
Ijms 24 01261 g007 550
Figure 7. Heatmap of GRF in M. dodecandrum. Note: different colors represent different FPKM values; the color gradient from blue to red represents increasing expression level.
Figure 7. Heatmap of GRF in M. dodecandrum. Note: different colors represent different FPKM values; the color gradient from blue to red represents increasing expression level.
Ijms 24 01261 g007
Ijms 24 01261 g008 550
Figure 8. Relative expression for MdGRFs in different organs. Note: abscissa represents different organs; ordinate represents value of relative expression.
Figure 8. Relative expression for MdGRFs in different organs. Note: abscissa represents different organs; ordinate represents value of relative expression.
Ijms 24 01261 g008
Ijms 24 01261 g009 550
Figure 9. Subcellular localizations of MdGRF19 in tobacco leaf. Scale bars = 10 µm. 35S::GFP was used as the empty control.
Figure 9. Subcellular localizations of MdGRF19 in tobacco leaf. Scale bars = 10 µm. 35S::GFP was used as the empty control.
Ijms 24 01261 g009
Table
Table 1. Physicochemical properties of GRF gene family proteins in M. dodecandrum.
Table 1. Physicochemical properties of GRF gene family proteins in M. dodecandrum.
GeneGene IDNumber of Amino AcidsMolecular Weight (Average)PIInstability IndexGrand Average of Hydropathicity (GRAVY)Subcellular Localization
MdGRF1DR00006644148,554.347.3454.19−0.693Nucleus
MdGRF2DR00087540545,832.019.1961.19−0.76Nucleus
MdGRF3DR00154638643,184.828.6957.13−0.816Nucleus
MdGRF4DR02281739944,121.329.357.08−0.809Nucleus
MdGRF5DR00948929231,752.637.244.85−0.475Nucleus
MdGRF6DR00558759263,117.058.1249.33−0.518Nucleus
MdGRF7DR01753038742,203.036.8561.5−0.674Nucleus
MdGRF8DR01616727729,999.537.7244.32−0.569Nucleus
MdGRF9DR02830047751,389.046.6256.85−0.623Nucleus
MdGRF10DR01320956760,022.766.4467.12−0.796Nucleus
MdGRF11DR02263043246,814.38.0245.27−0.542Nucleus
MdGRF12DR01766529833,057.798.1953.3−0.652Nucleus
MdGRF13DR03009143948,810.128.7969.76−0.859Nucleus
MdGRF14DR01255240044,738.518.9760.93−0.839Nucleus
MdGRF15DR01304230333,526.38.8953.52−0.706Nucleus
MdGRF16DR00676044349,686.16.9974.3−0.773Nucleus
MdGRF17DR01600535939,646.598.6754.35−0.585Nucleus
MdGRF18DR02204334536,736.868.849.51−0.481Nucleus
MdGRF19DR03422763768,659.678.0254.68−0.486Nucleus
MdGRF20DR03546855159,247.325.9459.39−0.662Nucleus
Table
Table 2. Selective pressure analysis of MdGRFs.
Table 2. Selective pressure analysis of MdGRFs.
Sequence 1Sequence 2KaKsKa/Ks
MdGRF1MdGRF110.1734933070.3988253330.43501075
MdGRF2MdGRF50.1324853950.8507839380.155721552
MdGRF2MdGRF60.0638556430.2808411540.227372814
MdGRF3MdGRF50.0770352290.214314820.359448912
MdGRF3MdGRF60.1514476690.7979592290.189793743
MdGRF6MdGRF50.1418882610.9078513870.156290185
MdGRF8MdGRF40.2159522730.9821056160.219887015
MdGRF10MdGRF70.1398520660.5601014870.249690582
MdGRF11MdGRF90.3508664430.9762543540.359400643
MdGRF14MdGRF100.1437916420.58274950.246746916
MdGRF14MdGRF70.0539951830.1419927530.380267179
MdGRF15MdGRF40.2279325140.8607714030.264800287
MdGRF15MdGRF80.0479742420.2084513250.230146014
MdGRF16MdGRF120.2584054560.9412497040.274534435
MdGRF16MdGRF170.12238470.2729334510.44840491
MdGRF17MdGRF120.3254391830.9695762270.335650951
MdGRF18MdGRF130.1928969220.8196189270.235349521
MdGRF19MdGRF130.0614811260.2010147170.305853855
MdGRF19MdGRF180.1815140360.766745530.236733087
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.

Share and Cite

MDPI and ACS Style

Huang, J.; Chen, G.-Z.; Ahmad, S.; Hao, Y.; Chen, J.-L.; Zhou, Y.-Z.; Lan, S.-R.; Liu, Z.-J.; Peng, D.-H. Genome-Wide Identification and Characterization of the GRF Gene Family in Melastoma dodecandrum. Int. J. Mol. Sci. 2023, 24, 1261. https://doi.org/10.3390/ijms24021261

AMA Style

Huang J, Chen G-Z, Ahmad S, Hao Y, Chen J-L, Zhou Y-Z, Lan S-R, Liu Z-J, Peng D-H. Genome-Wide Identification and Characterization of the GRF Gene Family in Melastoma dodecandrum. International Journal of Molecular Sciences. 2023; 24(2):1261. https://doi.org/10.3390/ijms24021261

Chicago/Turabian Style

Huang, Jie, Gui-Zhen Chen, Sagheer Ahmad, Yang Hao, Jin-Liao Chen, Yu-Zhen Zhou, Si-Ren Lan, Zhong-Jian Liu, and Dong-Hui Peng. 2023. "Genome-Wide Identification and Characterization of the GRF Gene Family in Melastoma dodecandrum" International Journal of Molecular Sciences 24, no. 2: 1261. https://doi.org/10.3390/ijms24021261

APA Style

Huang, J., Chen, G. -Z., Ahmad, S., Hao, Y., Chen, J. -L., Zhou, Y. -Z., Lan, S. -R., Liu, Z. -J., & Peng, D. -H. (2023). Genome-Wide Identification and Characterization of the GRF Gene Family in Melastoma dodecandrum. International Journal of Molecular Sciences, 24(2), 1261. https://doi.org/10.3390/ijms24021261

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

Article Metrics

Back to TopTop