Molecular mechanisms underlying floral trait formation in Phalaenopsis orchids

Abstract

Phalaenopsis orchids are one of the most important ornamental crops, prized for their beautiful flowers and long flowering phase. Hundreds of commercially available cultivars display a remarkable range of variation in key horticultural traits, including inflorescence type, floral size, and color patterning. While most current cultivars have been developed through cross-breeding or mutation breeding, genetic homogenization has become a growing concern. This is largely due to extensive hybridization among existing cultivars, which are predominantly derived from a limited number of parental species. Additionally, trait linkage in Phal. can hinder the integration of desirable characteristics in progeny. Therefore, there is an urgent need to decipher the genetic programs governing key horticultural traits to facilitate both conventional and molecular breeding. Despite significant research efforts, progress has been hampered by several resource limitations. These include a scarcity of high-quality genome assemblies, the lack of stable genetic transformation systems, and insufficient materials for molecular biology studies—a challenge exacerbated by the plant’s relatively long life cycle. Consequently, the molecular mechanisms underlying the formation and diversity of most important horticultural traits in Phal. orchids remain largely unexplored. This review summarizes recent research advances, with a primary focus on the key floral traits in Phal. orchids, including inflorescence type, flowering time, floral organ organization, color patterning, size, longevity, scent, organ shape, cuticle production, and wax biosynthesis. Furthermore, we offer perspectives on future research directions aimed at elucidating the genetic basis for the remarkable diversity of these traits and advancing molecular breeding in Phal. orchids.

Acknowledgements

We are grateful to the three anonymous reviewers for their insightful comments, which have significantly enhanced the quality of this manuscript. We thank Mr Junjie Liao for providing pictures of P. ‘Xiaohuizi’ and P. tetraspis ‘Purple Dream.’ We thank Prof. Jue Ruan from Agricultural Genomics Institute at Shenzhen, Chinese Academy of Agricultural Sciences for partial funding support. This work was supported by the National Natural Science Foundation of China (32370241), the Scientific Research Foundation of Kunpeng Institute of Modern Agriculture at Foshan (KIMAQD2022003), and the collaborative project (HXQS2024009 from Prof. Jue Ruan) to P.W.; Project supported by the Foundation for Innovative Research Groups of the National Natural Science Foundation of China (32221001) to H.K.; Chinese Universities Scientific Fund (2452024391) to R.Z.; National Key R&D Program of China (2023YFD1600505) to R.J.; the National Natural Science Foundation of China (32101584) and the Characteristic Innovation Project by the Guangdong Provincial Department of Education (2023KTSCX134) to T.X.

Author contributions

P.W., H.K., R.J., and R.Z. conceived and designed the outline of the manuscript. F.W. and P.W. made the figures. F.W., P.W., X.Z., and A.W.S. wrote the manuscript with inputs from all authors. All authors read and approved the final manuscript.

Conflicts of interest statement

The authors declare that they have no conflict of interests.

References

[1] Liu Z, Wang J, Olaf G. et al. The Genus Phalaenopsis in the World. Beijing: Higher Education Press. 2021:

[2] Hsu C-C, Chen H-H, Chen W-H. Chapter 23 Phalaenopsis. In: Johan Van Huylenbroech, Editor. Ornamental Crops. Springer International Publishing: Cham, 2018,567-626

[3] Zhou S, Jiang L, Guan S. et al. Expression profiles of five FT - like genes and functional analysis of PhFT-1 in a Phalaenopsis hybrid. Electron J Biotechnol. 2018; 31:75-83

[4] Jang S. Functional characterization of PhapLEAFY, a FLORI- CAULA / LEAFY ortholog in Phalaenopsis aphrodite. Plant Cell Physiol. 2015; 56:2234-47

[5] Chen D, Guo B, Hexige S. et al. SQUA -like genes in the orchid Phalaenopsis are expressed in both vegetative and reproduc- tive tissues. Planta. 2007; 226:369-80

[6] Ke Y-T, Lin K-F, Gu C-H. et al. Molecular characterization and expression profile of PaCOL1, a CONSTANS -like gene in Pha- laenopsis orchid. Plants. 2020; 9:68

[7] Koh KW, Lee S-H, Chen H-K. et al. Phalaenopsis flowering locus VE regulates floral organ maturation. Plant Cell Rep. 2018; 37:467-82

[8] Jiang L, Ren Y, Luo H. et al. Identification and functional analysis of J-domain proteins involved in determining flow- ering time in Phalaenopsis orchids. Plant Physiol Biochem. 2025; 226:110081

[9] Semiarti E. et al. Overexpression of PaFT Gene in the Wild Orchid Phalaenopsis amabilis (L.) Blume. The 5th international conference on mathematics and natural sciences. Bandung, Indonesia. 2015:090005

[10] Tsai W-C, Kuoh C-S, Chuang M-H. et al. Four DEF -like MADS box genes displayed distinct floral morphogenetic roles in Phalaenopsis orchid. Plant Cell Physiol. 2004; 45: 831-44

[11] Pan Z, Chen YY, du JS. et al. Flower development of Pha- laenopsis orchid involves functionally divergent SEPALLATA - like genes. New Phytol. 2014; 202:1024-42

[12] Acri-Nunes-Miranda R, Mondragón-Palomino M. Expressionof paralogous SEP -, FUL -, AG - and STK -like MADS-BOX genes in wild-type and peloric Phalaenopsis flowers. Front Plant Sci. 2014; 5:76

[13] Xu Q, Yang Z, Jia Y. et al. PeNAC67-PeKAN2-PeSCL23 and B- class MADS-BOX transcription factors synergistically regulate the specialization process from petal to lip in Phalaenopsis equestris. Mol Hortic. 2024; 4:15

[14] Su C, Chen WC, Lee AY. et al. A modified ABCDE model of flow- ering in orchids based on gene expression profiling studies of the moth orchid Phalaenopsis aphrodite. PLoS One. 2013; 8: e80462

[15] Lin Y-F, Chen YY, Hsiao YY. et al. Genome-wide identification and characterization of TCP genes involved in ovule develop- ment of Phalaenopsis equestris. J Exp Bot. 2016; 67:5051-66

[16] Hsu H-F, Hsu WH, Lee YI. et al. Model for perianth formation in orchids. Nat Plants. 2015; 1:15046

[17] Wang R, Mao C, Ming F. PeMYB4L interacts with PeMYC4 to reg- ulate anthocyanin biosynthesis in Phalaenopsis orchid. Plant Sci. 2022; 324:111423

[18] Hsieh M-H, Pan ZJ, Lai PH. et al. Virus-induced gene silenc- ing unravels multiple transcription factors involved in floral growth and development in Phalaenopsis orchids. J Exp Bot. 2013; 64:3869-84

[19] Hsu C-C, Su C-J, Jeng M-F. et al. A HORT1 retrotransposon insertion in the PeMYB11 promoter causes harlequin/black flowers in Phalaenopsis orchids. Plant Physiol. 2019; 180: 1535-48

[20] Zhao A, Cui Z, Li T. et al. mRNA and miRNA expression analysis reveal the regulation for flower spot patterning in Phalaenop- sis ‘Panda’. Int J Mol Sci. 2019; 20:4250

[21] Fu Z, Wang L, Shang H. et al. An R3-MYB gene of Phalaenopsis, MYBx1, represses anthocyanin accumulation. Plant Growth Regul. 2019; 88:129-38

[22] Hsiao Y, Jeng MF, Tsai WC. et al. A novel homodimeric geranyl diphosphate synthase from the orchid Phalaenopsis bellina lacking a DD( X ) 2-4 D motif. Plant J. 2008; 55:719-33

[23] Huang H, Kuo YW, Chuang YC. et al. Terpene synthase-b and terpene synthase-e/f genes produce monoterpenes for Phalaenopsis bellina floral scent. Front Plant Sci. 2021; 12: 700958

[24] Chang Y, Huang LM, Kuo XZ. et al. PbABCG1 and PbABCG2 transporters are required for the emission of floral monoter- penes in Phalaenopsis bellina. Plant J. 2023; 114:279-92

[25] Chuang Y-C, Hung Y-C, Tsai W-C. et al. PbbHLH4 regulates floral monoterpene biosynthesis in Phalaenopsis orchids. J Exp Bot. 2018; 69:4363-77

[26] Chuang Y-C, Hung YC, Hsu CY. et al. A dual repeat cis-element determines expression of GERANYL DIPHOSPHATE SYNTHASE for monoterpene production in Phalaenopsis orchids. Front Plant Sci. 2018; 9:765

[27] Hsu H-F, Li Y-C, Shen Y-H. et al.PaWOX3 and PaWOX3B regulate flower number and the lip symmetry of Phalaenopsis. Plant Cell Physiol. 2024; 65:1328-43

[28] Meng Y, He G, Zhang Q. etal. PeTB1/2 and PeSLR1 mediate the mechanism of PePIN1b involved in axillary bud dormancy in Phalaenopsis. Sci Hortic. 2025; 341:113949

[29] Lai P-H, Huang LM, Pan ZJ. et al. PeERF1, a SHINE-Like transcription factor, is involved in nanoridge development on lip epidermis of Phalaenopsis flowers. Front Plant Sci. 2020; 10:1709

[30] Mao W-T, Hsu W-H, Song J-L. et al. The HD-ZIP II gene PaHAT14 increases cuticle deposition by downregulating ERF gene PaERF105 in Phalaenopsis. Plant Cell Physiol. 2024; 65: 1751-68

[31] Lu H-C, Lam SH, Zhang D. et al. R2R3-MYB genes coordinate conical cell development and cuticular wax biosynthesis in Phalaenopsis aphrodite. Plant Physiol. 2022; 188:318-31

[32] Tsai W-C, Lee PF, Chen HI. et al. PeMADS6, a GLOBOSA/ PISTILLATA -like gene in Phalaenopsis equestris involved in petaloid formation, and correlated with flower longevity and ovary development. Plant Cell Physiol. 2005; 46:1125-39

[33] Chen W-H, Jiang Z-Y, Hsu H-F. et al. Silencing of FOREVER YOUNG FLOWER -like genes from Phalaenopsis orchids pro- motes flower senescence and abscission. Plant Cell Physiol. 2021; 62:111-24

[34] Chen W, Hsu W, Hsu H. et al. A tetraspanin gene regulating auxin response and affecting orchid perianth size and various plant developmental processes. Plant Direct. 2019; 3:e00157

[35] Wu Y-J, Chen SY, Hsu FC. et al. PeCIN8 expression correlates with flower size and resistance to yellow leaf disease in Pha- laenopsis orchids. BMC Plant Biol. 2023; 23:545

[36] Su W-R, Chen WS, Koshioka M. etal. Changes in gibberellin lev-els in the flowering shoot of Phalaenopsis hybrida under high temperature conditions when flower development is blocked. Plant Physiol Biochem. 2001; 39:45-50

[37] Lee HB, Im NH, An SK. et al. Changes of growth and inflorescence initiation by exogenous gibberellic acid 3 and 6-benzylaminopurine application in Phalaenopsis orchids. Agronomy. 2021; 11:196

[38] Blanchard MG, Runkle ES. Temperature during the day, but not during the night, controls flowering of Phalaenopsis orchids. J Exp Bot. 2006; 57:4043-9

[39] Huang J-Z, Lin CP, Cheng TC. et al. The genome and transcrip- tome of Phalaenopsis yield insights into floral organ develop- ment and flowering regulation. PeerJ. 2016; 4:e2017

[40] Blanchard MG, Runkle ES. Benzyladenine promotes flowering in Doritaenopsis and Phalaenopsis orchids. J Plant Growth Regul. 2008; 27:141-50

[41] Wang W-Y, Chen W-S, Chen W-H. et al. Influence of abscisic acid on flowering in Phalaenopsis hybrida. Plant Physiol Biochem. 2002; 40:97-100

[42] Ding L, Wang Y, Yu H. Overexpression of DOSOC1, an ortholog of Arabidopsis SOC1, promotes flowering in the orchid Dendrobium Chao Parya Smile. Plant Cell Physiol. 2013; 54:595-608

[43] Theißen G. Development of floral organ identity: stories from the MADS house. Curr Opin Plant Biol. 2001; 4:75-85

[44] Mondragón-Palomino M, Theißen G. MADS about the evolu- tion of orchid flowers. Trends Plant Sci. 2008; 13:51-9

[45] Pan Z-J, Cheng CC, Tsai WC. et al. The duplicated B-class MADS-BOX genes display dualistic characters in orchid flo- ral organ identity and growth. Plant Cell Physiol. 2011; 52: 1515-31

[46] Hsu H-F, Chen W-H, Shen Y-H. et al. Multifunctional evolu- tion of B and AGL6 MADS box genes in orchids. Nat Commun. 2021; 12:902

[47] Tsai CC, Chiang YC, Huang SC. et al. Molecular phylogeny of Phalaenopsis Blume (Orchidaceae) on the basis of plastid and nuclear DNA. Plant Syst Evol. 2010; 288:77-98

[48] Yao X, Zhang W, Duan X. et al. The making of elaborate petals in Nigella through developmental repatterning. New Phytol. 2019; 223:385-96

[49] Tsai W-C, Chen H-H. The orchid MADS-box genes controlling floral morphogenesis. Sci World J. 2006; 6:1933-44

[50] Huang J-Z, Lin CP, Cheng TC. et al. A de novo floral transcrip- tome reveals clues into Phalaenopsis orchid flower develop- ment. PLoS One. 2015; 10:e0123474

[51] Chen YH, Tsai YJ, Huang JZ. et al. Transcription analysis of peloric mutants of Phalaenopsis orchids derived from tissue culture. Cell Res. 2005; 15:639-57

[52] Hsieh M-H, Lu HC, Pan ZJ. et al. Optimizing virus-induced gene silencing efficiency with Cymbidium mosaic virus in Pha- laenopsis flower. Plant Sci. 2013;201-202:25-41

[53] Luo D, Carpenter R, Vincent C. et al. Origin of floral asymmetry in antirrhinum. Nature. 1996; 383:794-9

[54] Luo D, Carpenter R, Copsey L. et al. Control of organ asymme- try in flowers of Antirrhinum. Cell. 1999; 99:367-76

[55] Galego L, Almeida J. Role of DIVARICATA in the control of dorsoventral asymmetry in Antirrhinum flowers. Genes Dev. 2002; 16:880-91

[56] Preston JC, Hileman LC. Developmental genetics of floral symmetry evolution. Trends Plant Sci. 2009; 14:147-54

[57] Raimundo J, Sobral R, Bailey P. et al. A subcellular tug of war involving three MYB-like proteins underlies a molecu- lar antagonism in Antirrhinum flower asymmetry. Plant J. 2013; 75:527-38

[58] Preston JC, Hileman LC. Parallel evolution of TCP and B-class genes in Commelinaceae flower bilateral symmetry. EvoDevo. 2012; 3:6

[59] Bartlett ME, Williams SK, Taylor Z. et al. The maize PI / GLO ortholog Zmm16 / sterile tassel silky ear1 interacts with the zygomorphy and sex determination pathways in flower devel- opment. Plant Cell. 2015; 27:3081-98

[60] Valoroso MC, Sobral R, Saccone G. et al. Evolutionary conser- vation of the orchid MYB transcription factors DIV, RAD, and DRIF. Front Plant Sci. 2019; 10:1359

[61] Su S, Shao X, Zhu C. et al. Transcriptome-wide analysis reveals the origin of peloria in Chinese Cymbidium (Cymbid- ium sinense ). Plant Cell Physiol. 2018; 59:2064-74

[62] Hsu C-C, Chen Y-Y, Tsai W-C. et al. Three R2R3-MYB transcrip- tion factors regulate distinct floral pigmentation patterning in Phalaenopsis spp. Plant Physiol. 2015; 168:175-91

[63] Shang Y, Venail J, Mackay S. et al. The molecular basis for venation patterning of pigmentation and its effect on pollina- tor attraction in flowers of Antirrhinum. New Phytol. 2011; 189: 602-15

[64] Ding B, Patterson EL, Holalu SV. et al. Two MYB proteins in a self-organizing activator-inhibitor system produce spotted pigmentation patterns. Curr Biol. 2020; 30:802-814.e8

[65] Lu T-W, Chen W-H, Chen P-Y. et al. Correction: perturbation of periodic spot-generation balance leads to diversified pigmen- tation patterning of harlequin Phalaenopsis orchids: in silico prediction. BMC Plant Biol. 2024; 24:78266. Thathsarani YKDD, Tikirikumari JAT, Priyadarshan AIS. et al. Phytochemical evaluation of selected Phalaenopsis cultivars. Adv Hortic Sci. 2025; 39:125

[66] Liang C-Y, Rengasamy KP, Huang LM. et al. Assessment of violet-blue color formation in Phalaenopsis orchids. BMC Plant Biol. 2020; 20:212

[67] Zhao X, Zhang Y, Long T. et al. Regulation mechanism of plant pigments biosynthesis: anthocyanins, carotenoids, and betalains. Metabolites. 2022; 12:871

[68] Chen W-H, Hsu C-Y, Cheng H-Y. et al. Downregulation of puta- tive UDP-glucose: flavonoid 3-O-glucosyltransferase gene alters flower coloring in Phalaenopsis. Plant Cell Rep. 2011; 30: 1007-17

[69] Lou Y, Zhang Q, Xu Q. et al. PhCHS5 and PhF3 ′ 5′H genes over-expression in petunia (Petunia hybrida ) and Phalaenop- sis (Phalaenopsis aphrodite ) regulate flower color and branch number. Plants. 2023; 12:2204

[70] Xu W, Dubos C, Lepiniec L. Transcriptional control of flavonoid biosynthesis by MYB–bHLH–WDR complexes. Trends Plant Sci. 2015; 20:176-85

[71] Hsiao Y-Y, Tsai WC, Kuoh CS. et al. Comparison of transcripts in Phalaenopsis bellina and Phalaenopsis equestris (Orchi- daceae) flowers to deduce monoterpene biosynthesis path- way. BMC Plant Biol. 2006; 6:14

[72] Yeh C-H, Tsai WY, Chiang HM. et al. Headspace solid- phase microextraction analysis of volatile components in Phalaenopsis Nobby’s Pacific Sunset. Molecules. 2014; 19:14080-93

[73] Chen J, Zhu X, Zheng R. et al. Orchestrating of native Pha- laenopsis flower scents lighted the way through artificial selective breeding partiality in the current resource utiliza- tion. Ind Crop Prod. 2024; 217:118850

[74] Awano K, Honda T, Ogawa T. et al. Volatile components of Phalaenopsis schilleriana Rehb. f. Flavour Fragr J. 1997; 12:341-4

[75] Mus AA, Gansau JA, Kumar VS. et al. The variation of volatile compounds emitted from aromatic orchid (Phalaenopsis bel- lina ) at different timing and flowering stages. Plant Omics. 2020; 13:78-85

[76] Chuang Y-C, Lee M-C, Chang Y-L. et al. Diurnal regulation of the floral scent emission by light and circadian rhythm in the Phalaenopsis orchids. Bot Stud. 2017; 58:50

[77] Pramanik D, Spaans M, Kranenburg T. et al. Inflorescence lignification of natural species and horticultural hybrids of Phalaenopsis orchids. Sci Hortic. 2022; 295:110845

[78] Tsai S-S, Chang Y-CA. Plant maturity affects flowering ability and flower quality in Phalaenopsis, focusing on their relation- ship to carbon-to-nitrogen ratio. HortScience. 2022; 57:191-6

[79] Wang Y-T. Potassium nutrition affects Phalaenopsis growth and flowering. HortScience. 2007; 42:1563-7

[80] Xiao W, Li Z, Chen H. et al. Visualization of micromorphology of petal epidermal features of waxy and velvety flowers in Phalaenopsis. ScienceAsia. 2020; 46:657

[81] Porat R, Halevy AH, Serek M. et al. An increase in ethylene sensitivity following pollination is the initial event trigger- ing an increase in ethylene production and enhanced senes- cence of Phalaenopsis orchid flowers. Physiol Plant. 1995; 93: 778-84

[82] Zhang W, Li J, Wang D. etal. Study on ploidy identification and fertility correlation of Phalaenopsis germplasm resources. J Nucl Agric Sci. 2024; 38:1012-23

[83] Sevilleno SS, Cabahug-Braza RA, An HR. et al. Morphological and cytogenetic evaluation of Phalaenopsis cultivars. Hortic Environ Biotechnol. 2025; 66:1421-35

[84] Ho T-T, Kwon A-R, Yoon Y-J. et al. Endoreduplication level affects flower size and development by increasing cell size in Phalaenopsis and Doritaenopsis. Acta Physiol Plant. 2016; 38:190

[85] Guan Y, Zhang Q, Zhang T. et al. Transcriptome analysis reveals the mechanisms underlying petal growth during the flower opening process in Phalaenopsis orchids. BMC Plant Biol. 2025; 25:696

[86] Guan Y, Wong CE, Zhang Q. et al. Sizing up beauty: mech- anisms of petal size regulation in ornamental plants. Plant Physiol. 2025;198:kiaf198

[87] Pramanik D, Dorst N, Meesters N. et al. Evolution and devel- opment of three highly specialized floral structures of bee- pollinated Phalaenopsis species. EvoDevo. 2020; 11:16

[88] Cai J, Liu X, Vanneste K. et al. The genome sequence of the orchid Phalaenopsis equestris. Nat Genet. 2015; 47: 65-72

[89] Chao Y, Chen WC, Chen CY. et al. Chromosome-levelassembly, genetic and physical mapping of Phalaenopsis aphrodite genome provides new insights into species adaptation and resources for orchid breeding. Plant Biotechnol J. 2018; 16:2027-41

[90] Zhang D, Zhao XW, Li YY. et al. Advances and prospects of orchid research and industrialization. Hortic Res. 2022;9: uhac220

[91] Enoki S, Takahara Y. Applications of biotechnological approaches in the product and breeding of Phalaenopsis orchids. In: Sarwar Khan M,ed. Tropical Plant Species and Technological Interventions for Improvement. London: IntechOpen, 2023,

[92] Zhou Y, Zhang Z, Bao Z. et al. Graph pangenome captures missing heritability and empowers tomato breeding. Nature. 2022; 606:527-34

[93] Liu Z, Wang N, Su Y. et al. Grapevine pangenome facil- itates trait genetics and genomic breeding. Nat Genet. 2024; 56:2804-14

[94] Wang K, Shi L, Liang X. et al. The gene TaWOX5 overcomes genotype dependency in wheat genetic transformation. Nat Plants. 2022; 8:110-7

[95] Yang W, Zhai H, Wu F. et al. Peptide REF1 is a local wound signal promoting plant regeneration. Cell. 2024; 187: 3024-3038.e14