Identification and Expression Analysis of the β-Carotene Hydroxylase Gene Family of Oncidium hybridum

doi:10.16420/j.issn.0513-353x.2025-0745

Acta Horticulturae Sinica ›› 2026, Vol. 53 ›› Issue (6): 1637-1651.doi: 10.16420/j.issn.0513-353x.2025-0745

• Genetic & Breeding · Germplasm Resources · Molecular Biology • Previous Articles Next Articles

Identification and Expression Analysis of the β-Carotene Hydroxylase Gene Family of Oncidium hybridum

CHEN Yan¹, ZHENG Jinxuan², LUO Yuanhua¹, CHEN Yiquan¹, FANG Nengyan¹, ZHONG Huaiqin¹^,^*()

¹ Institute of Crop Sciences， Fujian Academy of Agricultural Sciences（Fujian Germplasm Resources Center）/Fujian Engineering Research Center for Characteristic Floriculture， Fuzhou 350013， China
² Institute of Horticultural Biotechnology， Fujian Agriculture and Forestry University， Fuzhou 350002， China

Received:2025-12-21 Revised:2026-03-19 Online:2026-06-24 Published:2026-06-24
Contact: ZHONG Huaiqin

Abstract

Abstract:

β-Carotene Hydroxylase（BCH）is a key rate limiting enzyme in the carotenoid biosynthesis pathway. Based on the genome data of Oncidium hybridum，the BCH family was identified. The gene structure，chromosome localization，physicochemical properties，phylogenetic and collinear relationships of the BCH family were analyzed. The BCHs expression patterns were analyzed at different stages of flower color change in O. hybridum‘XM-1’and among different Oncidium cultivars. The results showed that nine BCH members were identified in the genome of O. hybridum，distributed on eight chromosomes，and there were fragment duplication events. The phylogenetic tree indicated that OnBCH mainly cluster in two branches. The upstream promoter region contained various regulatory elements involved in light signals，stress responses，growth and development，and hormone regulatory elements. The qRT-PCR results showed that the expression levels of OnBCH1/3/4 were significantly down-regulated during the half flowering stage（S2，yellow）to the full flowering stage（S4，white）of‘XM-1’. This trend was consistent with the changes in lutein，zeaxanthin，and β-cryptoxanthin content. It was speculated that OnBCH1/3/4 play an important role in the color dynamics of‘XM-1’. There were significant differences in carotenoid content among 12 cultivars of yellow series O. hybridum. Based on qRT-PCR results，it was speculated that genes OnBCH1/3/4，as well as genes OnBCH6/7/8/9，have similarities in regulating carotenoid mediated coloration function in O. hybridum. An OnBCH transcription factor regulatory network was constructed. It was speculated that the MYB-related transcription factor evm.model.Chr084.732 and the BCH3 regulatory module play important roles in the carotenoid-mediated flower color change process of O. hybridum.

Key words: Oncidium hybridum, β-carotene hydroxylase, BCH gene family, carotenoid

CHEN Yan, ZHENG Jinxuan, LUO Yuanhua, CHEN Yiquan, FANG Nengyan, ZHONG Huaiqin. Identification and Expression Analysis of the β-Carotene Hydroxylase Gene Family of Oncidium hybridum[J]. Acta Horticulturae Sinica, 2026, 53(6): 1637-1651.

Figures/Tables 10

Fig. 1 Phenotype of flower color changes of Oncidium‘XM-1’during flowering（A）and 12 yellow cultivars of Oncidium hybridum（B）

Table 1 The sequences of qRT-PCR primers

引物名称Primer name	上游引物（5′-3′）Forward primer	下游引物（5′-3′）Reverse primer
OnBCH1	GCCTTCGCTATGTCCTCTT	ACCGAGCAACCAATGGAT
OnBCH2	TCGCCAAAGTTGCTATGC	GCTTCTTCTTCTTCTCAGCG
OnBCH3	GCCTTTGCTATGTCCTCTTC	GCTACACCGAACAACCAAT
OnBCH4	CGTCCCGTATTTCCAATG	ATTCTTCCATCCCTCCCA
OnBCH5	ACGAGCAACAGGAGAAGAGC	CGATGGCAGGAGAAGAGAAC
OnBCH6	TTGTCTATCTGTGGGTGCTG	GCCTGTGATGCGATTCAT
OnBCH7	GCCATCATCGCAGTCTACT	GTCCCAAGCATTTCCGTAT
OnBCH8	CCACCATCTGCTTTCTCAT	AGTAGACTGCGATGATGGC
OnBCH9	GCAGCCGTTATGTCCAGTT	CAGCACCCACAGATAGACAA
OnActin	GGAGAAACTTGCTTATGTGGC	CGTAATGACTTGACCATCCG

Table 2 Basic parameters analysis of OnBCH family

基因ID Gene ID	基因名称 Gene name	氨基酸数 Number of amino acids	分子量/Da Molecular weight	等电点 pI	不稳定系数 Instability coefficient	脂肪族指数 Aliphatic index	亲水性 Gravy	亚细胞定位 Subcellular localization
evm.model.Chr045.315	OnBCH1	410	46 174.80	6.99	43.42	68.56	-0.257	叶绿体 Chloroplast
evm.model.Chr046.218	OnBCH2	190	21 704.58	10.63	46.02	92.00	0.476	细胞质Cytoplasmic
evm.model.Chr047.194	OnBCH3	386	43 497.33	9.77	47.67	80.67	-0.084	叶绿体 Chloroplast
evm.model.Chr048.203	OnBCH4	171	19 273.15	7.25	45.75	78.77	0.084	叶绿体 Chloroplast
evm.model.Chr085.590	OnBCH5	597	66 523.38	8.71	66.65	76.33	-0.322	线粒体Mitochondrial
evm.model.Chr086.400	OnBCH6	280	31 010.88	6.93	70.10	84.43	0.096	叶绿体 Chloroplast
evm.model.Chr087.477	OnBCH7	304	33 796.12	4.69	60.51	85.76	0.020	叶绿体 Chloroplast
evm.model.Chr087.567	OnBCH8	300	33 436.60	5.22	66.71	84.93	-0.026	叶绿体 Chloroplast
evm.model.Chr088.437	OnBCH9	304	33 700.91	9.87	70.15	86.38	0.027	叶绿体 Chloroplast

Fig. 2 Chromosomal localization（A）and replication events（B）of the OnBCH family members in Oncidium hybridum

Fig. 3 Conservative motif and gene structure analysis of the BCH family proteins in Oncidium hybridum

Fig. 4 Analysis of promoter elements of BCH family genes in Oncidium hybridum

Fig. 5 Expression analysis of OnBCH family genes of Oncidium‘XM-1’ S1：Initial flowering stage；S2：Half flowering stage；S3：Flowering stage；S4：Full flowering stage. Different lowercase letters indicate significant differences among different samples at the P < 0.05 level

Fig. 6 Carotenoid content determination of different cultivars of Oncidium hybridum BY：Oncidium Gower Ramsey‘White Jade’；MT：Oncidium Sweet Sugar‘Wide’；SY：Oncidium pusillum；JH：Oncidium Gower Ramsey‘Jinhui’；NML：Oncidium Sweet Sugar‘Lemon Drop’；MS：Oncidium Tropic Breeze‘Ever Glade’；XS：Oncidium Jairak Fragrance；LH：Willsonara Dandy‘Sundance’；CF：Odontocidium Spring Maple；KK：Oncidium Space Race‘CoCo’；XJS：Oncidium Gower Ramsey‘Sunkist’；HJL：Oncidium‘Golden Spirit’. Different lowercase letters indicate significant differences among different samples at the P < 0.05 level. The same below

Fig. 7 Expression levels of OnBCH family members in different cultivars of Oncidium hybridum

Fig. 8 Predictive regulatory networks of BCH family genes and transcription factors in Oncidium hybridum

References 36

[1]	Cai J, Liu X, Vanneste K, Proost S, Tsai W C, Liu K W, Chen L J, He Y, Xu Q, Bian C. 2015. The genome sequence of the orchid Phalaenopsis equestris. Nat Genet, 47 (1):65-72.
[2]	Castillo R, Fernández J A, Gómez-Gómez L. 2005. Implications of carotenoid biosynthetic genes in apocarotenoid formation during the stigma development of Crocus sativus and its closer relatives. Plant Physiol, 139 (2):674-689.
[3]	Cazzonelli C I, Pogson B J. 2010. Source to sink:regulation of carotenoid biosynthesis in plants. Trends in Plant Science, 15 (5):266-274.
[4]	Chen C, Wu Y, Li J, Wang X, Zeng Z, Xu J, Liu Y, Feng J, Chen H, He Y, Xia R. 2023. TBtools-II:A“one for all,all for one”bioinformatics platform for biological big-data mining. Mol Plant, 16 (11):1733-1742.
[5]	Chiou C Y, Pan H A, Chuang Y N, Yeh K W. 2010. Differential expression of carotenoid-related genes determines diversified carotenoid coloration in floral tissues of Oncidium cultivars. Planta, 232 (4):937-948.
[6]	Davison P A, Hunter C N, Horton P. 2002. Overexpression of beta-carotene hydroxylase enhances stress tolerance in Arabidopsis. Nature, 418 (6894):203-206.
[7]	Diretto G, Welsch R, Tavazza R, Mourgues F, Pizzichini D, Beyer P, Giuliano G. 2007. Silencing of beta-carotene hydroxylase increases total carotenoid and beta-carotene levels in potato tubers. BMC Plant Biol,7:11.
[8]	Fan R H, Lin B, Fang N Y, Ye X X, Huang M L, Zhong H Q. 2020. Transcriptome-sequencing analyses reveal flower color formation in Strelitzia reginae. Biologia Plantarum,64:717-724.
[9]	Fraser P D, Bramley P M. 2004. The biosynthesis and nutritional uses of carotenoids. Prog Lipid Res, 43 (3):228-265.
[10]	Galpaz N, Ronen G, Khalfa Z, Zamir D, Hirschberg J. 2006. A chromoplast-specific carotenoid biosynthesis pathway is revealed by cloning of the tomato white-flower locus. Plant Cell, 18 (8):1947-1960.
[11]	He Jingjuan, Fan Yanping. 2022. Progress in composition and metabolic regulation of carotenoids related to floral color. Acta Horticulturae Sinica, 49 (5):1162-1172. (in Chinese)
	何静娟, 范燕萍. 2022. 观赏植物花色相关的类胡萝卜素组成及代谢调控研究进展. 园艺学报, 49 (5):1162-1172.
[12]	Huang Xinlei, Wang Yan, Zhang Hui. 2019. Analysis of carotenoids compounds and their biosynthesis pathways in flowers of three Dendrobium species. Forest Res, 32 (5):107-113. (in Chinese)
	黄昕蕾, 王雁, 张辉. 2019. 3种石斛属植物类胡萝卜素成分及代谢途径分析. 林业科学研究, 32 (5):107-113.
[13]	Iijima L, Kishimoto S, Ohmiya A, Yagi M, Okamoto E, Miyahara T, Tsujimoto T, Ozeki Y, Uchiyama N, Hakamatsuka T, Kouno T,A. Cano E, Shimizu M, Nishihara M. 2020. Esterified carotenoids are synthesized in petals of carnation(Dianthus caryophyllus)and accumulate in differentiated chromoplasts. Sci Rep,10:15256.
[14]	Kim S H, Ahn Y O, Ahn M J, Lee H S, Kwak S S. 2012. Down-regulation of beta-carotene hydroxylase increases beta-carotene and total carotenoids enhancing salt stress tolerance in transgenic cultured cells of sweetpotato. Phytochemistry,74:69-78.
[15]	Kumar S, Stecher G, Li M, Knyaz C, Tamura K. 2018. MEGA X:molecular evolutionary genetics analysis across computing platforms. Mol Biol Evol, 35 (6):1547-1549.
[16]	Li T, Liu J X, Deng Y J, Duan A Q, Liu H, Zhuang F Y, Xiong A S. 2023. Differential hydroxylation efficiency of the two non-heme carotene hydroxylases:DcBCH1,rather than DcBCH2,plays a major role in carrot taproot. Hortic Res, 9 (1):967-979.
[17]	Li Tong. 2022. Functional analysis of carrot DcBCH and DcCCD4 in carotenoid metabolism[Ph. D. Dissertation]. Nanjing: Nanjing Agricultural University. (in Chinese)
	李彤. 2022. 胡萝卜DcBCH和DcCCD4在类胡萝卜素代谢中的功能分析[博士论文]. 南京: 南京农业大学.
[18]	Luo Y, Chen Y, Fang N, Kong L, Lin R, Chen Y, Fan R, Zhong H, Huang M, Ye X. 2024. Multiomics analysis reveals the involvement of OnDIVARICATA 3 in controlling dynamic flower coloring of Oncidium hybridum. Plant Physiol Biochem,217:109277.
[19]	Ma X, Zheng B, Ma Y, Xu W, Wu H, Wang S. 2018. Carotenoid accumulation and expression of carotenoid biosynthesis genes in mango flesh during fruit development and ripening. Scientia Horticulturae,237:201-206.
[20]	Pons E, Alquézar B, Rodríguez A, Martorell P, Genovés S, Ramón D, Rodrigo M J, Zacarías L, Pe A L. 2014. Metabolic engineering of β-carotene in orange fruit increases its in vivo antioxidant properties. Plant Biotechnol J, 12 (1):17-27.
[21]	Prakash A, Jeffryes M, Bateman A, Finn R D. 2017. The HMMER web server for protein sequence similarity search. Current Protocols in Bioinformatics, 60 (1):3.15.1-3.15.23.
[22]	Pu X, Li Z, Tian Y, Gao R, Hao L, Hu Y, He C, Sun W, Xu M, Peters R, Peer Y, Xu Z, Song J. 2020. The honeysuckle genome provides insight into the molecular mechanism of carotenoid metabolism underlying dynamic flower coloration. New Phytol, 227 (3):930-943.
[23]	Song J, Sun B, Chen C, Ning Z, Zhang S, Cai Y, Zheng X, Cao B, Chen G, Jin D. 2023. An R-R-type MYB transcription factor promotes non-climacteric pepper fruit carotenoid pigment biosynthesis. The Plant J, 115 (3):724-741.
[24]	Sridhar D, Carlo R, Patrizia P, Riccardo A, Florence B, Bilal C, Giovanni G. 2002. Metabolic engineering of xanthophyll content in tomato fruits. Febs Letters, 519 (1-3):30-34.
[25]	Sun Q, He Z, Wei R, Zhang Y, Ye J, Chai L, Xie Z, Guo W, Xu J, Cheng Y. 2024. The transcriptional regulatory module CsHB5-CsbZIP44 positively regulates abscisic acid-mediated carotenoid biosynthesis in citrus(Citrus spp.). Plant Biotechnol J, 22 (3):722-737.
[26]	Tian L. 2003. Functional analysis of beta- and epsilon-ring carotenoid hydroxylases in Arabidopsis. The Plant Cell, 15 (6):1320-1332.
[27]	Tian L, Dellapenna D. 2001. Characterization of a second carotenoid β-hydroxylase gene from Arabidopsis and its relationship to the LUT1 locus. Plant Mol Biol, 47 (3):379-388.
[28]	Vallabhaneni R, Gallagher C E, Licciardello N, Cuttriss A J, Quinlan R F, Wurtzel E T. 2009. Metabolite sorting of a germplasm collection reveals the Hydroxylase3 Locus as a new target for maize provitamin a biofortification. Plant Physiol, 151 (3):1635-1645.
[29]	Wang H M, To K Y, Lai H M, Jeng S T. 2016. Modification of flower colour by suppressing β-ring carotene hydroxylase genes in Oncidium. Plant Biol, 18 (2):220-229.
[30]	Xia H, Zhou Y, Lin Z, Guo Y, Liu X, Wang T, Wang J, Deng H, Lin L, Deng Q. 2022. Characterization and functional validation of beta-carotene hydroxylase AcBCH genes in Actinidia chinensis. Hortic Res,9:uhac063.
[31]	Xiong L, Schumaker K S, Zhu J K. 2002. Cell signaling during cold,drought,and salt stress. The Plant Cell,14 (Suppl):S165-S183.
[32]	Yu B, Lydiate D J, Schafer U A, Hannoufa A. 2007. Characterization of a beta-carotene hydroxylase of Adonis aestivalis and its expression in Arabidopsis thaliana. Planta, 226 (1):181-192.
[33]	Zhang D, Zhao X W, Li Y Y, Ke S J, Yin W L, Lan S, Liu Z J. 2022a. Advances and prospects of orchid research and industrialization. Hortic Res,9:uhac220.
[34]	Zhang X, Lin S, Peng D, Wu Q, Liao X, Xiang K, Wang Z, Tembrock L R, Bendahmane M, Bao M. 2022b. Integrated multi-omic data and analyses reveal the pathways underlying key ornamental traits in carnation flowers. Plant Biotechnol J, 20 (6):1182-1196.
[35]	Zhang Y, Jin J, Zhu S, Sun Q, Zhang Y, Xie Z, Ye J, Deng X. 2023. Citrus β-carotene hydroxylase 2(BCH_(2))participates in xanthophyll synthesis by catalyzing the hydroxylation of β-carotene and compensates for BCH1 in citrus carotenoid metabolism. Hortic Res, 10 (3):45-56.
[36]	Zhou Yuanjie. 2022. Functional validation of β-carotene hydroxylase gene in kiwifruit[M. D. Dissertation]. Ya’an: Sichuan Agricultural University. (in Chinese)
	周源洁. 2022. 猕猴桃β-胡萝卜素羟化酶基因的功能验证[硕士论文]. 雅安: 四川农业大学.

Identification and Expression Analysis of the β-Carotene Hydroxylase Gene Family of Oncidium hybridum

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