R2R3-MYB genes coordinate conical cell development and cuticular wax biosynthesis in Phalaenopsis aphrodite.

Petals of the monocot Phalaenopsis aphrodite (Orchidaceae) possess conical epidermal cells on their adaxial surfaces, and a large amount of cuticular wax is deposited on them to serve as a primary barrier against biotic and abiotic stresses. It has been widely reported that subgroup 9A members of the R2R3-MYB gene family, MIXTA and MIXTA-like in eudicots, act to regulate the differentiation of conical epidermal cells. However, the molecular pathways underlying conical epidermal cell development and cuticular wax biosynthesis in monocot petals remain unclear. Here, we characterized two subgroup 9A R2R3-MYB genes, PaMYB9A1 and PaMYB9A2 (PaMYB9A1/2), from P. aphrodite through the transient overexpression of their coding sequences and corresponding chimeric repressors in developing petals. We showed that PaMYB9A1/2 function to coordinate conical epidermal cell development and cuticular wax biosynthesis. In addition, we identified putative targets of PaMYB9A1/2 through comparative transcriptome analyses, revealing that PaMYB9A1/2 acts to regulate the expression of cell wall-associated and wax biosynthetic genes. Furthermore, a chemical composition analysis of cuticular wax showed that even-chain n-alkanes and odd-chain primary alcohols are the main chemical constituents of cuticular wax deposited on petals, which is inconsistent with the well-known biosynthetic pathways of cuticular wax, implying a distinct biosynthetic pathway occurring in P. aphrodite flowers. These results reveal that the function of subgroup 9A R2R3-MYB family genes in regulating the differentiation of epidermal cells is largely conserved in monocots and dicots. Furthermore, both PaMYB9A1/2 have evolved additional functions controlling the biosynthesis of cuticular wax.

Genome-Wide Identification Analysis of the R2R3-MYB Transcription Factor Family in Cymbidium sinense for Insights into Drought Stress Responses.

Cymbidium sinense represents a distinctive Orchidaceae plant that is more tolerant than other terrestrial orchids. Studies have shown that many members of the MYB transcription factor (TF) family, especially the R2R3-MYB subfamily, are responsive to drought stress. This study identified 103 CsMYBs; phylogenetic analysis classified these genes into 22 subgroups with Arabidopsis thaliana. Structural analysis showed that most CsMYB genes contained the same motifs, three exons and two introns, and showed a helix-turn-helix 3D structure in each R repeat. However, the members of subgroup 22 contained only one exon and no intron. Collinear analysis revealed that C. sinense had more orthologous R2R3-MYB genes with wheat than A. thaliana and rice. Ka/Ks ratios indicated that most CsMYB genes were under purifying negative selection pressure. Cis-acting elements analysis revealed that drought-related elements were mainly focused on subgroups 4, 8, 18, 20, 21, and 22, and Mol015419 (S20) contained the most. The transcriptome analysis results showed that expression patterns of most CsMYB genes were upregulated in leaves in response to slight drought stress and downregulated in roots. Among them, members in S8 and S20 significantly responded to drought stress in C. sinense. In addition, S14 and S17 also participated in these responses, and nine genes were selected for the real-time reverse transcription quantitative PCR (RT-qPCR) experiment. The results were roughly consistent with the transcriptome. Our results, thus, provide an important contribution to understanding the role of CsMYBs in stress-related metabolic processes.

Characteristics and Comparative Analysis of Seven Complete Plastomes of Trichoglottis s.l. (Aeridinae, Orchidaceae).

Trichoglottis exhibits a range of rich variations in colors and shapes of flower and is a valuable ornamental orchid genus. The genus Trichoglottis has been expanded by the inclusion of Staurochilus, but this Trichoglottis sensu lato (s.l.) was recovered as a non-monophyletic genus based on molecular sequences from one or a few DNA regions. Here, we present phylogenomic data sets, incorporating complete plastome sequences from seven species (including five species sequenced in this study) of Trichoglottis s.l. (including two species formerly treated as Staurochilus), to compare plastome structure and to reconstruct the phylogenetic relationships of this genus. The seven plastomes possessed the typical quadripartite structure of angiosperms and ranged from 149,402 bp to 149,841 bp with a GC content of 36.6-36.7%. These plastomes contain 120 genes, which comprise 74 protein-coding genes, 38 tRNA genes, and 8 rRNA genes, all ndh genes were pseudogenized or lost. A total of 98 (T. philippinensis) to 134 (T. ionosma) SSRs and 33 (T. subviolacea) to 46 (T. ionosma) long repeats were detected. The consistent and robust phylogenetic relationships of Trichoglottis were established using a total of 25 plastid genomes from the Aeridinae subtribe. The genus Trichoglottis s.l. was strongly supported as a monophyletic group, and two species formerly treated as Staurochilus were revealed as successively basal lineages. In addition, five mutational hotspots (trnNGUU-rpl32, trnLUAA, trnSGCU-trnGUCC, rbcL-accD, and trnTGGU-psbD) were identified based on the ranking of PI values. Our research indicates that plastome data is a valuable source for molecular identification and evolutionary studies of Trichoglottis and its related genera.

Apostasia Mitochondrial Genome Analysis and Monocot Mitochondria Phylogenomics.

Apostasia shenzhenica belongs to the subfamily Apostasioideae and is a primitive group located at the base of the Orchidaceae phylogenetic tree. However, the A. shenzhenica mitochondrial genome (mitogenome) is still unexplored, and the phylogenetic relationships between monocots mitogenomes remain unexplored. In this study, we discussed the genetic diversity of A. shenzhenica and the phylogenetic relationships within its monocotyledon mitogenome. We sequenced and assembled the complete mitogenome of A. shenzhenica, resulting in a circular mitochondrial draft of 672,872 bp, with an average read coverage of 122× and a GC content of 44.4%. A. shenzhenica mitogenome contained 36 protein-coding genes, 16 tRNAs, two rRNAs, and two copies of nad4L. Repeat sequence analysis revealed a large number of medium and small repeats, accounting for 1.28% of the mitogenome sequence. Selection pressure analysis indicated high mitogenome conservation in related species. RNA editing identified 416 sites in the protein-coding region. Furthermore, we found 44 chloroplast genomic DNA fragments that were transferred from the chloroplast to the mitogenome of A. shenzhenica, with five plastid-derived genes remaining intact in the mitogenome. Finally, the phylogenetic analysis of the mitogenomes from A. shenzhenica and 28 other monocots showed that the evolution and classification of most monocots were well determined. These findings enrich the genetic resources of orchids and provide valuable information on the taxonomic classification and molecular evolution of monocots.

Comprehensive phylogenetic analyses of Orchidaceae using nuclear genes and evolutionary insights into epiphytism.

Orchidaceae (with >28,000 orchid species) are one of the two largest plant families, with economically and ecologically important species, and occupy global and diverse niches with primary distribution in rainforests. Among orchids, 70% grow on other plants as epiphytes; epiphytes contribute up to ~50% of the plant diversity in rainforests and provide food and shelter for diverse animals and microbes, thereby contributing to the health of these ecosystems. Orchids account for over two-thirds of vascular epiphytes and provide an excellent model for studying evolution of epiphytism. Extensive phylogenetic studies of Orchidaceae and subgroups have ;been crucial for understanding relationships among many orchid lineages, although some uncertainties remain. For example, in the largest subfamily Epidendroideae with nearly all epiphytic orchids, relationships among some tribes and many subtribes are still controversial, hampering evolutionary analyses of epiphytism. Here we obtained 1,450 low-copy nuclear genes from 610 orchid species, including 431 with newly generated transcriptomes, and used them for the reconstruction of robust Orchidaceae phylogenetic trees with highly supported placements of tribes and subtribes. We also provide generally well-supported phylogenetic placements of 131 genera and 437 species that were not sampled by previous plastid and nuclear phylogenomic studies. Molecular clock analyses estimated the Orchidaceae origin at ~132 million years ago (Ma) and divergences of most subtribes from 52 to 29 Ma. Character reconstruction supports at least 14 parallel origins of epiphytism; one such origin was placed at the most recent common ancestor of ~95% of epiphytic orchids and linked to modern rainforests. Ten occurrences of rapid increase in the diversification rate were detected within Epidendroideae near and after the K-Pg boundary, contributing to ~80% of the Orchidaceae diversity. This study provides a robust and the largest family-wide Orchidaceae nuclear phylogenetic tree thus far and new insights into the evolution of epiphytism in vascular plants.

The Euscaphis japonica genome and the evolution of malvids.

Malvids is one of the largest clades of rosids, includes 58 families and exhibits remarkable morphological and ecological diversity. Here, we report a high-quality chromosome-level genome assembly for Euscaphis japonica, an early-diverging species within malvids. Genome-based phylogenetic analysis suggests that the unstable phylogenetic position of E. japonica may result from incomplete lineage sorting and hybridization event during the diversification of the ancestral population of malvids. Euscaphis japonica experienced two polyploidization events: the ancient whole genome triplication event shared with most eudicots (commonly known as the γ event) and a more recent whole genome duplication event, unique to E. japonica. By resequencing 101 samples from 11 populations, we speculate that the temperature has led to the differentiation of the evergreen and deciduous of E. japonica and the completely different population histories of these two groups. In total, 1012 candidate positively selected genes in the evergreen were detected, some of which are involved in flower and fruit development. We found that reddening and dehiscence of the E. japonica pericarp and long fruit-hanging time promoted the reproduction of E. japonica populations, and revealed the expression patterns of genes related to fruit reddening, dehiscence and abscission. The key genes involved in pentacyclic triterpene synthesis in E. japonica were identified, and different expression patterns of these genes may contribute to pentacyclic triterpene diversification. Our work sheds light on the evolution of E. japonica and malvids, particularly on the diversification of E. japonica and the genetic basis for their fruit dehiscence and abscission.

OrchidBase 5.0: updates of the orchid genome knowledgebase.

Containing the largest number of species, the orchid family provides not only materials for studying plant evolution and environmental adaptation, but economically and culturally important ornamental plants for human society. Previously, we collected genome and transcriptome information of Dendrobium catenatum, Phalaenopsis equestris, and Apostasia shenzhenica which belong to two different subfamilies of Orchidaceae, and developed user-friendly tools to explore the orchid genetic sequences in the OrchidBase 4.0. The OrchidBase 4.0 offers the opportunity for plant science community to compare orchid genomes and transcriptomes and retrieve orchid sequences for further study.In the year 2022, two whole-genome sequences of Orchidoideae species, Platanthera zijinensis and Platanthera guangdongensis, were de novo sequenced, assembled and analyzed. In addition, systemic transcriptomes from these two species were also established. Therefore, we included these datasets to develop the new version of OrchidBase 5.0. In addition, three new functions including synteny, gene order, and miRNA information were also developed for orchid genome comparisons and miRNA characterization.OrchidBase 5.0 extended the genetic information to three orchid subfamilies (including five orchid species) and provided new tools for orchid researchers to analyze orchid genomes and transcriptomes. The online resources can be accessed at https://cosbi.ee.ncku.edu.tw/orchidbase5/.

OrchidBase 4.0: a database for orchid genomics and molecular biology.

BACKGROUND: The Orchid family is the largest families of the monocotyledons and an economically important ornamental plant worldwide. Given the pivotal role of this plant to humans, botanical researchers and breeding communities should have access to valuable genomic and transcriptomic information of this plant. Previously, we established OrchidBase, which contains expressed sequence tags (ESTs) from different tissues and developmental stages of Phalaenopsis as well as biotic and abiotic stress-treated Phalaenopsis. The database includes floral transcriptomic sequences from 10 orchid species across all the five subfamilies of Orchidaceae. DESCRIPTION: Recently, the whole-genome sequences of Apostasia shenzhenica, Dendrobium catenatum, and Phalaenopsis equestris were de novo assembled and analyzed. These datasets were used to develop OrchidBase 4.0, including genomic and transcriptomic data for these three orchid species. OrchidBase 4.0 offers information for gene annotation, gene expression with fragments per kilobase of transcript per millions mapped reads (FPKM), KEGG pathways and BLAST search. In addition, assembled genome sequences and location of genes and miRNAs could be visualized by the genome browser. The online resources in OrchidBase 4.0 can be accessed by browsing or using BLAST. Users can also download the assembled scaffold sequences and the predicted gene and protein sequences of these three orchid species. CONCLUSIONS: OrchidBase 4.0 is the first database that contain the whole-genome sequences and annotations of multiple orchid species. OrchidBase 4.0 is available at http://orchidbase.itps.ncku.edu.tw/.

The genome of Cymbidium sinense revealed the evolution of orchid traits.

The Orchidaceae is of economic and ecological importance and constitutes ˜10% of all seed plant species. Here, we report a genome physical map for Cymbidium sinense, a well-known species belonging to genus Cymbidium that has thousands of natural variation varieties of flower organs, flower and leaf colours and also referred as the King of Fragrance, which make it arose into a unique cultural symbol in China. The high-quality chromosome-scale genome assembly was 3.52 Gb in size, 29 638 protein-coding genes were predicted, and evidence for whole-genome duplication shared with other orchids was provided. Marked amplification of cytochrome- and photosystem-related genes was observed, which was consistent with the shade tolerance and dark green leaves of C. sinense. Extensive duplication of MADS-box genes, and the resulting subfunctional and expressional differentiation, was associated with regulation of species-specific flower traits, including wild-type and mutant-type floral patterning, seasonal flowering and ecological adaption. CsSEP4 was originally found to positively regulate gynostemium development. The CsSVP genes and their interaction proteins CsAP1 and CsSOC1 were significantly expanded and involved in the regulation of low-temperature-dependent flowering. Important genetic clues to the colourful leaf traits, purple-black flowers and volatile trait in C. sinense were also found. The results provide new insights into the molecular mechanisms of important phenotypic traits of Cymbidium and its evolution and serve as a powerful platform for future evolutionary studies and molecular breeding of orchids.

Molecular insights into self-incompatibility systems: From evolution to breeding.

Plants have evolved diverse self-incompatibility (SI) systems for outcrossing. Since Darwin's time, considerable progress has been made toward elucidating this unrivaled reproductive innovation. Recent advances in interdisciplinary studies and applications of biotechnology have given rise to major breakthroughs in understanding the molecular pathways that lead to SI, particularly the strikingly different SI mechanisms that operate in Solanaceae, Papaveraceae, Brassicaceae, and Primulaceae. These best-understood SI systems, together with discoveries in other "nonmodel" SI taxa such as Poaceae, suggest a complex evolutionary trajectory of SI, with multiple independent origins and frequent and irreversible losses. Extensive exploration of self-/nonself-discrimination signaling cascades has revealed a comprehensive catalog of male and female identity genes and modifier factors that control SI. These findings also enable the characterization, validation, and manipulation of SI-related factors for crop improvement, helping to address the challenges associated with development of inbred lines. Here, we review current knowledge about the evolution of SI systems, summarize key achievements in the molecular basis of pollen‒pistil interactions, discuss potential prospects for breeding of SI crops, and raise several unresolved questions that require further investigation.

Genome-wide characterization and expression profiling of the HD-ZIP gene family in Acoraceae under salinity and cold stress.

The Homeodomain-Leucine Zipper (HD-ZIP) transcription factors play a pivotal role in governing various aspects of plant growth, development, and responses to abiotic stress. Despite the well-established importance of HD-ZIPs in many plants, their functions in Acoraceae, the basal lineage of monocots, remain largely unexplored. Using recently published whole-genome data, we identified 137 putative HD-ZIPs in two Acoraceae species, Acorus gramineus and Acorus calamus. These HD-ZIP genes were further classified into four subfamilies (I, II, III, IV) based on phylogenetic and conserved motif analyses, showcasing notable variations in exon-intron patterns among different subfamilies. Two microRNAs, miR165/166, were found to specifically target HD-ZIP III genes with highly conserved binding sites. Most cis-acting elements identified in the promoter regions of Acoraceae HD-ZIPs are involved in modulating light and phytohormone responsiveness. Furthermore, our study revealed an independent duplication event in Ac. calamus and a one-to-multiple correspondence between HD-ZIP genes of Ac. calamus and Ac. gramineus. Expression profiles obtained from qRT-PCR demonstrated that HD-ZIP I genes are strongly induced by salinity stress, while HD-ZIP II members have contrasting stress responses in two species. HD-ZIP III and IV genes show greater sensitivity in stress-bearing roots. Taken together, these findings contribute valuable insights into the roles of HD-ZIP genes in stress adaptation and plant resilience in basal monocots, illuminating their multifaceted roles in plant growth, development, and response to abiotic stress.

Genomic Resequencing Unravels the Genetic Basis of Domestication, Expansion, and Trait Improvement in Morus Atropurpurea.

Mulberry is an economically important plant in the sericulture industry and traditional medicine. However, the genetic and evolutionary history of mulberry remains largely unknown. Here, this work presents the chromosome-level genome assembly of Morus atropurpurea (M. atropurpurea), originating from south China. Population genomic analysis using 425 mulberry accessions reveal that cultivated mulberry is classified into two species, M. atropurpurea and M. alba, which may have originated from two different mulberry progenitors and have independent and parallel domestication in north and south China, respectively. Extensive gene flow is revealed between different mulberry populations, contributing to genetic diversity in modern hybrid cultivars. This work also identifies the genetic architecture of the flowering time and leaf size. In addition, the genomic structure and evolution of sex-determining regions are identified. This study significantly advances the understanding of the genetic basis and domestication history of mulberry in the north and south, and provides valuable molecular markers of desirable traits for mulberry breeding.

Diploid and tetraploid genomes of Acorus and the evolution of monocots.

Monocots are a major taxon within flowering plants, have unique morphological traits, and show an extraordinary diversity in lifestyle. To improve our understanding of monocot origin and evolution, we generate chromosome-level reference genomes of the diploid Acorus gramineus and the tetraploid Ac. calamus, the only two accepted species from the family Acoraceae, which form a sister lineage to all other monocots. Comparing the genomes of Ac. gramineus and Ac. calamus, we suggest that Ac. gramineus is not a potential diploid progenitor of Ac. calamus, and Ac. calamus is an allotetraploid with two subgenomes A, and B, presenting asymmetric evolution and B subgenome dominance. Both the diploid genome of Ac. gramineus and the subgenomes A and B of Ac. calamus show clear evidence of whole-genome duplication (WGD), but Acoraceae does not seem to share an older WGD that is shared by most other monocots. We reconstruct an ancestral monocot karyotype and gene toolkit, and discuss scenarios that explain the complex history of the Acorus genome. Our analyses show that the ancestors of monocots exhibit mosaic genomic features, likely important for that appeared in early monocot evolution, providing fundamental insights into the origin, evolution, and diversification of monocots.

Genome-Wide Identification and Expression Pattern Analysis of KNOX Gene Family in Orchidaceae.

The establishment of lateral organs and subsequent plant architecture involves factors intrinsic to the stem apical meristem (SAM) from which they are derived. KNOTTED1-LIKE HOMEOBOX (KNOX) genes are a family of plant-specific homeobox transcription factors that especially act in determining stem cell fate in SAM. Although KNOXs have been studied in many land plants for decades, there is a dearth of knowledge on KNOX's role in Orchidaceae, the largest and most diverse lineage of flowering plants. In this study, a total of 32 putative KNOX genes were identified in the genomes of five orchid species and further designated into two classes (Class I and Class II) based on phylogenetic relationships. Sequence analysis showed that most orchid KNOX proteins retain four conserved domains (KNOX1, KNOX2, ELK, and Homeobox_KN). Comparative analysis of gene structure showed that the exon-intron structure is conserved in the same clade but most orchids exhibited longer intron, which may be a unique feature of Orchidaceae. Cis-elements identified in the promoter region of orchid KNOXs were found mostly enriched in a function of light responsiveness, followed by MeJA and ABA responsiveness, indicative of their roles in modulating light and phytohormones. Collinear analysis unraveled a one-to-one correspondence among KNOXs in orchids, and all KNOX genes experienced strong purifying selection, indicating the conservation of this gene family has been reinforced across the Orchidaceae lineage. Expression profiles based on transcriptomic data and real-time reverse transcription-quantitative PCR (RT-qPCR) revealed a stem-specific expression of KNOX Class I genes and a broader expression pattern of Class II genes. Taken together, our results provided a comprehensive analysis to uncover the underlying function of KNOX genes in Orchidaceae.

Advances and prospects of orchid research and industrialization.

Orchidaceae is one of the largest, most diverse families in angiosperms with significant ecological and economical values. Orchids have long fascinated scientists by their complex life histories, exquisite floral morphology and pollination syndromes that exhibit exclusive specializations, more than any other plants on Earth. These intrinsic factors together with human influences also make it a keystone group in biodiversity conservation. The advent of sequencing technologies and transgenic techniques represents a quantum leap in orchid research, enabling molecular approaches to be employed to resolve the historically interesting puzzles in orchid basic and applied biology. To date, 16 different orchid genomes covering four subfamilies (Apostasioideae, Vanilloideae, Epidendroideae, and Orchidoideae) have been released. These genome projects have given rise to massive data that greatly empowers the studies pertaining to key innovations and evolutionary mechanisms for the breadth of orchid species. The extensive exploration of transcriptomics, comparative genomics, and recent advances in gene engineering have linked important traits of orchids with a multiplicity of gene families and their regulating networks, providing great potential for genetic enhancement and improvement. In this review, we summarize the progress and achievement in fundamental research and industrialized application of orchids with a particular focus on molecular tools, and make future prospects of orchid molecular breeding and post-genomic research, providing a comprehensive assemblage of state of the art knowledge in orchid research and industrialization.

Genome-wide identification and expression analysis of the GRAS gene family in Dendrobium chrysotoxum.

The GRAS gene family encodes transcription factors that participate in plant growth and development phases. They are crucial in regulating light signal transduction, plant hormone (e.g. gibberellin) signaling, meristem growth, root radial development, response to abiotic stress, etc. However, little is known about the features and functions of GRAS genes in Orchidaceae, the largest and most diverse angiosperm lineage. In this study, genome-wide analysis of the GRAS gene family was conducted in Dendrobium chrysotoxum (Epidendroideae, Orchidaceae) to investigate its physicochemical properties, phylogenetic relationships, gene structure, and expression patterns under abiotic stress in orchids. Forty-six DchGRAS genes were identified from the D. chrysotoxum genome and divided into ten subfamilies according to their phylogenetic relationships. Sequence analysis showed that most DchGRAS proteins contained conserved VHIID and SAW domains. Gene structure analysis showed that intronless genes accounted for approximately 70% of the DchGRAS genes, the gene structures of the same subfamily were the same, and the conserved motifs were also similar. The Ka/Ks ratios of 12 pairs of DchGRAS genes were all less than 1, indicating that DchGRAS genes underwent negative selection. The results of cis-acting element analysis showed that the 46 DchGRAS genes contained a large number of hormone-regulated and light-responsive elements as well as environmental stress-related elements. In addition, the real-time reverse transcription quantitative PCR (RT-qPCR) experimental results showed significant differences in the expression levels of 12 genes under high temperature, drought and salt treatment, among which two members of the LISCL subfamily (DchGRAS13 and DchGRAS15) were most sensitive to stress. Taken together, this paper provides insights into the regulatory roles of the GRAS gene family in orchids.

Genome-wide identification and analysis of anthocyanin synthesis-related R2R3-MYB genes in Cymbidium goeringii.

The MYB gene family plays a vital regulatory role in plant metabolism, stress response, and floral color. The R2R3-MYB gene family of C. goeringii was identified, and its expression was analyzed using bioinformatics in this article. The R2R3-MYB genes of Arabidopsis thaliana were used as a reference to determine 104 CgMYB genes and categorize them into 22 subfamilies. Exon/intron organizations and conserved motif analysis revealed that the majority of CgMYB genes were highly conserved, and chromosome localization and collinearity analysis provided evidence of tandem duplication and segmental duplication events, indicating the phenomenon of gene family expansion and contraction. The function of CgMYB genes was analyzed by cis-acting element and gene ontology (GO) enrichment. In addition, we selected CgMYB91 and CgMYB32 for RT-qPCR, suggesting that CgMYB91 and CgMYB32 are associated with anthocyanin formation. In short, this study provides a comprehensive and specific function of the R2R3-MYB transcription factors (TFs) in orchids.

Genome-wide analysis of the TCP gene family and their expression pattern in Cymbidium goeringii.

TCP gene family are specific transcription factors for plant, and considered to play an important role in development and growth. However, few related studies investigated the TCP gene trait and how it plays a role in growth and development of Orchidaceae. In this study, we obtained 14 TCP genes (CgTCPs) from the Spring Orchid Cymbidium goeringii genome. The classification results showed that 14 CgTCPs were mainly divided into two clades as follows: four PCF genes (Class I), nine CIN genes and one CYC gene (Class II). The sequence analysis showed that the TCP proteins of C. goeringii contain four conserved regions (basic Helix-Loop-Helix) in the TCP domain. The exon-intron structure varied in the clade according to a comparative investigation of the gene structure, and some genes had no introns. There are fewer CgTCP homologous gene pairs compared with Dendrobium catenatum and Phalaenopsis equestris, suggesting that the TCP genes in C. goeringii suffered more loss events. The majority of the cis-elements revealed to be enriched in the function of light responsiveness, followed by MeJA and ABA responsiveness, demonstrating their functions in regulating by light and phytohormones. The collinearity study revealed that the TCPs in D. catenatum, P. equestris and C. goeringii almost 1:1. The transcriptomic data and real-time reverse transcription-quantitative PCR (RT-qPCR) expression profiles showed that the flower-specific expression of the TCP class II genes (CgCIN2, CgCIN5 and CgCIN6) may be related to the regulation of florescence. Altogether, this study provides a comprehensive analysis uncovering the underlying function of TCP genes in Orchidaceae.

Genome-wide identification of YABBY genes in three Cymbidium species and expression patterns in C. ensifolium (Orchidaceae).

Members of the YABBY gene family play significant roles in lamina development in cotyledons, floral organs, and other lateral organs. The Orchidaceae family is one of the largest angiosperm groups. Some YABBYs have been reported in Orchidaceae. However, the function of YABBY genes in Cymbidium is currently unknown. In this study, 24 YABBY genes were identified in Cymbidium ensifolium, C. goeringii, and C. sinense. We analyzed the conserved domains and motifs, the phylogenetic relationships, chromosome distribution, collinear correlation, and cis-elements of these three species. We also analyzed expression patterns of C. ensifolium and C. goeringii. Phylogenetic relationships analysis indicated that 24 YABBY genes were clustered in four groups, INO, CRC/DL, YAB2, and YAB3/FIL. For most YABBY genes, the zinc finger domain was located near the N-terminus and the helix-loop-helix domain (YABBY domain) near the C-terminus. Chromosomal location analysis results suggested that only C. goeringii YABBY has tandem repeat genes. Almost all the YABBY genes displayed corresponding one-to-one relationships in the syntenic relationships analysis. Cis-elements analysis indicated that most elements were clustered in light-responsive elements, followed by MeJA-responsive elements. Expression patterns showed that YAB2 genes have high expression in floral organs. RT-qPCR analysis showed high expression of CeYAB3 in lip, petal, and in the gynostemium. CeCRC and CeYAB2.2 were highly expressed in gynostemium. These findings provide valuable information of YABBY genes in Cymbidium species and the function in Orchidaceae.

Deletion and tandem duplications of biosynthetic genes drive the diversity of triterpenoids in Aralia elata.

Araliaceae species produce various classes of triterpene and triterpenoid saponins, such as the oleanane-type triterpenoids in Aralia species and dammarane-type saponins in Panax, valued for their medicinal properties. The lack of genome sequences of Panax relatives has hindered mechanistic insight into the divergence of triterpene saponins in Araliaceae. Here, we report a chromosome-level genome of Aralia elata with a total length of 1.05 Gb. The loss of 12 exons in the dammarenediol synthase (DDS)-encoding gene in A. elata after divergence from Panax might have caused the lack of dammarane-type saponin production, and a complementation assay shows that overexpression of the PgDDS gene from Panax ginseng in callus of A. elata recovers the accumulation of dammarane-type saponins. Tandem duplication events of triterpene biosynthetic genes are common in the A. elata genome, especially for AeCYP72As, AeCSLMs, and AeUGT73s, which function as tailoring enzymes of oleanane-type saponins and aralosides. More than 13 aralosides are de novo synthesized in Saccharomyces cerevisiae by overexpression of these genes in combination. This study sheds light on the diversity of saponins biosynthetic pathway in Araliaceae and will facilitate heterologous bioproduction of aralosides.