甜瓜果皮颜色基因的鉴定与标记开发

doi:10.1016/j.jia.2024.11.004

Journal of Integrative Agriculture ›› 2025, Vol. 24 ›› Issue (7): 2589-2600.DOI: 10.1016/j.jia.2024.11.004

甜瓜果皮颜色基因的鉴定与标记开发

收稿日期:2024-02-22 修回日期:2024-11-04 接受日期:2024-09-27 出版日期:2025-07-20 发布日期:2025-06-17

Identification and molecular marker development for peel color gene in melon (Cucumis melo L.)

Jian Ma^1*^#, Guoliang Yuan¹^*, Xinyang Xu², Haijun Zhang¹, Yanhong Qiu¹, Congcong Li³^#, Huijun Zhang⁴^#

¹ Key Laboratory of Vegetable Biobreeding/Key Laboratory of Biology and Genetic Improvement of Horticultural Crops (North China) of Ministry of Agriculture and Rural Affairs, Beijing Vegetable Research Center (BVRC), Beijing Academy of Agriculture and Forestry Sciences, Beijing 100097, China
² Institute of Vegetables, Zhejiang Academy of Agricultural Sciences, Hangzhou 310021, China
³ Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, China
⁴ School of Life Science, Huaibei Normal University, Huaibei 235000, China

Received:2024-02-22 Revised:2024-11-04 Accepted:2024-09-27 Online:2025-07-20 Published:2025-06-17
About author: Jian Ma, E-mail: majian@nercv.org; Guoliang Yuan, E-mail: yuanguoliang@nercv.org #Correspondence Jian Ma, E-mail: majian@nercv.org; Congcong Li, E-mail: licongcong01@caas.cn; Huijun Zhang, E-mail: zhhuijun@126.com * These authors contributed equally to this study.
Supported by:
This work was supported by the Innovation and Development Program of Beijing Vegetable Research Center, China (KYCX202301), the Construction of Cucurbits Collaboration and Innovation Center, China (XTCX202301), the Youth Research Fund of Beijing Academy of Agriculture and Forestry Sciences, China (QNJJ202426), the National Natural Science Foundation of China (U21A20229 and 32102397), the Scientific Research Foundation of the Higher Education Institutions for Distinguished Young Scholars in Anhui Province, China (2022AH020037) and the Key Research and Development Projects of Anhui Province, China (2023z04020019).

摘要/Abstract

摘要：

果皮颜色是甜瓜的重要外观品质，会极大地影响着消费者的喜好。本研究中，我们利用灰绿色果皮甜瓜材料B8和白色果皮材料B15构建了一个深绿色果皮近等基因系NIL-G，随后以NIL-G和B15为亲本重新构建了次级F₂群体来研究果皮颜色的遗传模式，并进一步采用混池分离群体分析测序方法确定目标基因所在的物理位置。遗传分析表明：甜瓜成熟时的深绿色果皮性状由单显性基因控制。利用混池分离群体测序和分子标记将目的基因所在区间定位到4号染色体顶端263.7 kb区间内，该区间包含已报道的CmAPRR2基因。此外，进一步对NIL-G和B15中的CmAPRR2基因序列分析发现两者存在四个SNP变异，其中SNP.G614331A为G到A的突变，位于第6外显子和第6内含子的交界处，该突变导致B15中CmAPRR2基因转录本发生可变剪接，产生两个翻译提前终止的转录本CmAPRR2-A 和 CmAPRR2-B。我们也基于该SNP变异位点开发了一个KASP标记APRR2-G/A，该标记与构建的F₂群体中的果皮颜色表型共分离。另外，NIL-G中CmAPRR2的表达水平在果实各个生长阶段都高于B15。因此，我们推测CmAPRR2可能是控制甜瓜果实颜色的关键目的基因。总之，本研究鉴定到一个CmAPRR2的新等位基因，该等位变异导致甜瓜成熟时形成白色果皮。本研究为进一步研究甜瓜果皮颜色调控机制奠定了理论基础，并将来利用分子标记辅助选择改良甜瓜的果皮颜色提供了技术支撑。

Abstract:

Peel color is an important appearance quality of melons that significantly affects consumer preferences. In this study, a near-isogenic line NIL-G (dark green peel) was generated from B8 (grey-green peel) and B15 (white peel). The F₂ population constructed by crossing NIL-G and B15 was used to study the inheritance pattern of peel color, and bulked-segregant analysis sequencing (BSA-seq) was employed to identify the interval in which the target gene was located. Genetic analysis showed that a dominant gene controls the dark green peel trait at maturity. BSA-seq and molecular markers were used to localize the candidate gene in a 263.7 kb interval of chromosome 4, which contained the CmAPRR2 gene with known functions. Moreover, allelic sequence analysis revealed four SNP variations of the CmAPRR2 gene in B15, of which SNP.G614331A was located at the junction of the 6th exon and 6th intron. The G-to-A mutation caused alternative splicing of the transcript of CmAPRR2 in B15, generating two transcripts (CmAPRR2-A and CmAPRR2-B) with premature termination codons. Furthermore, the Kompetitive Allele Specific PCR (KASP) marker, APRR2-G/A, was developed based on this SNP and shown to co-segregate with the peel color phenotype in the F₂ population. Compared to white-peel B15, the expression level of CmAPRR2 in dark green peel NIL-G was higher at each growth stage. Therefore, CmAPRR2 may be the key gene controlling the fruit color of melons. This study identified a novel allelic variant of CmAPRR2 that leads to white peel formation in mature melons. We also provides a theoretical basis for further research on the gene regulatory mechanism of melon peel colors, which promotes using molecular marker-assisted selection to modify melon peel colors in the future.

Key words: melon , BSA-seq , mapping , CmAPRR2 , peel color , KASP marker

Jian Ma, Guoliang Yuan, Xinyang Xu, Haijun Zhang, Yanhong Qiu, Congcong Li, Huijun Zhang. 甜瓜果皮颜色基因的鉴定与标记开发[J]. Journal of Integrative Agriculture, 2025, 24(7): 2589-2600.

Jian Ma, Guoliang Yuan, Xinyang Xu, Haijun Zhang, Yanhong Qiu, Congcong Li, Huijun Zhang. Identification and molecular marker development for peel color gene in melon (Cucumis melo L.)[J]. Journal of Integrative Agriculture, 2025, 24(7): 2589-2600.

参考文献

Allan A C, Hellens R P, Laing W A. 2008. MYB transcription factors that colour our fruit. Trends in Plant Science, 13, 99–102.

Barash Y, Calarco J A, Gao W, Pan Q, Wang X, Shai O, Blencowe B J, Frey B J. 2010. Deciphering the splicing code. Nature, 465, 53–59.

Chen M, Ji M, Wen B, Liu L, Li S, Chen X, Gao D, Li L. 2016. GOLDEN 2-LIKE transcription factors of plants. Frontiers in Plant Science, 7, 1509.

Chen S, Zhou Y, Chen Y, Gu J. 2018. fastp: An ultra-fast all-in-one FASTQ preprocessor. Bioinformatics, 34, i884–i890.

Endl J, Achigan-Dako E G, Pandey A K, Monforte A J, Pico B, Schaefer H. 2018. Repeated domestication of melon (Cucumis melo) in Africa and Asia and a new close relative from India. American Journal of Botany, 105, 1662–1671.

Fang H, Wang P, Wang W, Peng J, Zheng J, Zhu G, Zhong C, Yu W. 2023. Fine mapping and identification of SmAPRR2 regulating rind color in eggplant (Solanum melongena L.). International Journal of Molecular Sciences, 24, 3059.

Feder A, Burger J, Gao S, Lewinsohn E, Katzir N, Schaffer A A, Meir A, Davidovich-Rikanati R, Portnoy V, Gal-On A, Fei Z, Kashi Y, Tadmor Y. 2015. A Kelch domain-containing F-box coding gene negatively regulates flavonoid accumulation in muskmelon. Plant Physiology, 169, 1714–1726.

Freilich S, Lev S, Gonda I, Reuveni E, Portnoy V, Oren E, Lohse M, Galpaz N, Bar E, Tzuri G, Wissotsky G, Meir A, Burger J, Tadmor Y, Schaffer A, Fei Z, Giovannoni J, Lewinsohn E, Katzir N. 2015. Systems approach for exploring the intricate associations between sweetness, color and aroma in melon fruits. BMC Plant Biology, 15, 71.

Gur A, Tzuri G, Meir A, Sa’ar U, Portnoy V, Katzir N, Schaffer A A, Li L, Burger J, Tadmor Y. 2017. Genome-wide linkage-disequilibrium mapping to the candidate gene level in melon (Cucumis melo). Scientific Reports, 7, 9770.

Hao N, Du Y, Li H, Wang C, Wang C, Gong S, Zhou S, Wu T. 2018. CsMYB36 is involved in the formation of yellow green peel in cucumber (Cucumis sativus L.). Theoretical and Applied Genetics, 131, 1659–1669.

Hassan M Z, Robin A H K, Rahim M A, Natarajan S, Kim H T, Park J I, Nou I S. 2018. Screening of melon genotypes identifies gummy stem blight resistance associated with Gsb1 resistant loci. Journal of Plant Biotechnology, 45, 217–227.

Hertle A P, García-Cerdán J G, Armbruster U, Shih R, Lee J J, Wong W, Niyogi K K. 2020. A Sec14 domain protein is required for photoautotrophic growth and chloroplast vesicle formation in Arabidopsis thaliana. Proceedings of the National Academy of Sciences of the United States of America, 117, 9101–9111.

Hill J T, Demarest B L, Bisgrove B W, Gorsi B, Su Y C, Yost H J. 2013. MMAPPR: Mutation mapping analysis pipeline for pooled RNA-seq. Genome Research, 23, 687–697.

Hosoda K, Imamura A, Katoh E, Hatta T, Tachiki M, Yamada H, Mizuno T, Yamazaki T. 2002. Molecular structure of the GARP family of plant Myb-related DNA binding motifs of the Arabidopsis response regulators. The Plant Cell, 14, 2015–2029.

Hu Z, Xu F, Guan L, Qian P, Liu Y, Zhang H, Huang Y, Hou S. 2014. The tetratricopeptide repeat-containing protein slow green1 is required for chloroplast development in Arabidopsis. Journal of Experimental Botany, 65, 1111–1123.

Huo Y, Zhang G, Yu W, Liu Z, Shen M, Zhao R, Hu S, Zheng X, Wang P, Yang Y. 2023. Forward genetic studies reveal LsAPRR2 as a key gene in regulating the green color of pericarp in bottle gourd (Lagenaria siceraria). Frontiers in Plant Science, 14, 1130669.

Jacob-Wilk D, Holland D, Goldschmidt E E, Riov J, Eyal Y. 1999. Chlorophyll breakdown by chlorophyllase: Isolation and functional expression of the Chlase1 gene from ethylene-treated Citrus fruit and its regulation during development. The Plant Journal, 20, 653–661.

Jeffrey C. 1980. A review of the cucurbitaceae. Botanical Journal of the Linnean Society, 81, 233–247.

Jeong H B, Jang S J, Kang M Y, Kim S, Kwon J K, Kang B C. 2020. Candidate gene analysis reveals that the fruit color locus C1 corresponds to PRR2 in pepper (Capsicum frutescens). Frontiers in Plant Science, 11, 399.

Jiao J, Liu H, Liu J, Cui M, Xu J, Meng H, Li Y, Chen S, Cheng Z. 2017. Identification and functional characterization of APRR2 controlling green immature fruit color in cucumber (Cucumis sativus L.). Plant Growth Regulation, 83, 233–243.

Jin H, Martin C. 1999. Multifunctionality and diversity within the plant MYB-gene family. Plant Molecular Biology, 41, 577–585.

Kerje T, Grum M. 2000. The origin of melon, Cucumis melo: A review of the literature. Acta Horticulturae, 510, 37–44.

Lewis B P, Green R E, Brenner S E. 2003. Evidence for the widespread coupling of alternative splicing and nonsense-mediated mRNA decay in humans. Proceedings of the National Academy of Sciences of the United States of America, 100, 189–192.

Li H, Handsaker B, Wysoker A, Fennell T, Ruan J, Homer N, Marth G, Abecasis G, Durbin R, 1000 Genome Project Data Processing Subgroup. 2009. The sequence alignment/map format and SAMtools. Bioinformatics, 25, 2078–2079.

Liu D, Liang J, Liu Q, Chen Y, Duan S, Sun D, Zhu H, Dou J, Niu H, Yang S, Sun S, Hu J, Yang L. 2025. The pseudo-type response regulator gene Clsc regulates rind stripe coloration in watermelon. Journal of Integrative Agriculture, 24, 147–160.

Liu H, Jiao J, Liang X, Liu J, Meng H, Chen S, Li Y, Cheng Z. 2016. Map-based cloning, identification and characterization of the w gene controlling white immature fruit color in cucumber (Cucumis sativus L.). Theoretical and Applied Genetics, 129, 1247–1256.

Liu L, Sun T, Liu X, Guo Y, Huang X, Gao P, Wang X. 2019. Genetic analysis and mapping of a striped rind gene (st3) in melon (Cucumis melo L.). Euphytica, 215, 20.

Lun Y, Wang X, Zhang C, Yang L, Gao D, Chen H, Huang S. 2016. CsYcf54 variant conferring light green coloration in cucumber. Euphytica, 208, 509–517.

Lv J, Fu Q, Lai Y, Zhou M, Wang H. 2018. Inheritance and gene mapping of spotted to non-spotted trait gene CmSp-1 in melon (Cucumis melo L. var. chinensis Pangalo). Molecular Breeding, 38, 105.

Lyu X, Xia Y, Wang C, Zhang K, Deng G, Shen Q, Gao W, Zhang M, Liao N, Ling J, Bo Y, Hu Z, Yang J, Zhang M. 2023. Pan-genome analysis sheds light on structural variation-based dissection of agronomic traits in melon crops. Plant Physiology, 193, 1330–1348.

Ma J, Li C, Zong M, Qiu Y, Liu Y, Huang Y, Xie Y, Zhang H, Wang J. 2022. CmFSI8/CmOFP13 encoding an OVATE family protein controls fruit shape in melon. Journal of Experimental Botany, 73, 1370–1384.

Ma J, Wang J. 2019. Development and application of specific molecular marker of the gynoecious gene g in melon. Journal of Plant Genetic Resources, 20, 1080–1086. (in Chinese)

Ma J, Yuan G, Zhang H, Qiu Y, Zong M, Zhu X, Zhang H. 2024. Development and application of a codominant marker of the melon rind colour gene CmAPRR2. Plant Breeding, 143, 276–283.

Ma L, Liu Z, Cheng Z, Gou J, Chen J, Yu W, Wang P. 2021. Identification and application of BhAPRR2 controlling peel colour in wax gourd (Benincasa hispida). Frontiers in Plant Science, 12, 716772.

Makino S, Kiba T, Imamura A, Hanaki N, Nakamura A, Suzuki T, Taniguchi M, Ueguchi C, Sugiyama T, Mizuno T. 2000. Genes encoding pseudo-response regulators: Insight into His-to-Asp phosphorelay and circadian rhythm in Arabidopsis thaliana. Plant Cell & Physiology, 41, 791–803.

Marquez Y, Brown J W, Simpson C, Barta A, Kalyna M. 2012. Transcriptome survey reveals increased complexity of the alternative splicing landscape in Arabidopsis. Genome Research, 22, 1184–1195.

Monforte A J, Oliver M, Gonzalo M J, Alvarez J M, Dolcet-Sanjuan R, Arús P. 2004. Identification of quantitative trait loci involved in fruit quality traits in melon (Cucumis melo L.). Theoretical and Applied Genetics, 108, 750–758.

Murray M G, Thompson W F. 1980. Rapid isolation of high molecular weight plant DNA. Nucleic Acids Research, 8, 4321–4325.

Nadakuduti S S, Holdsworth W L, Klein C L, Barry C S. 2014. KNOX genes influence a gradient of fruit chloroplast development through regulation of GOLDEN2-LIKE expression in tomato. The Plant Journal, 78, 1022–1033.

Nilsen T W, Graveley B R. 2010. Expansion of the eukaryotic proteome by alternative splicing. Nature, 463, 457–463.

Oren E, Tzuri G, Vexler L, Dafna A, Meir A, Sa’ar U, Faigenboim A, Kenigswald M, Portnoy V, Schaffer A, Levi A, Buckler E, Katzir N, Burger J, Tadmor Y, Gur A. 2019. The multi-allelic APRR2 gene is associated with fruit pigment accumulation in melon and watermelon. Journal of Experimental Botany, 70, 3781–3794.

Ou D. 2019. Genetic analysis and gene mapping for green peel color in melon (Cucumis melo L.). MSc thesis, Chinese Academy of Agricultural Sciences, Beijing, China. (in Chinese).

Ou D, Zhao G, He Y, Wang P, Xu Z, Kong W, Zhang J, Xu Y. 2019. Genetic analysis and gene mapping for melon rind color. Chinese Agricultural Science Bulletin, 35, 64–69. (in Chinese)

Pan Y, Bradley G, Pyke K, Ball G, Lu C, Fray R, Marshall A, Jayasuta S, Baxter C, van Wijk R, Boyden L, Cade R, Chapman N H, Fraser P D, Hodgman C, Seymour G B. 2013. Network inference analysis identifies an APRR2-like gene linked to pigment accumulation in tomato and pepper fruits. Plant Physiology, 161, 1476–1485.

Pei S, Liu T, Ren X, Li W, Chen C, Xie Z. 2021. Benchmarking variant callers in next-generation and third-generation sequencing analysis. Brief Bioinformatics, 22, bbaa148.

Pitrat M. 2013. Phenotypic diversity in wild and cultivated melons (Cucumis melo). Plant Biotechnology, 30, 273–278.

Pitrat M. 2016. Melon Genetic resources: Phenotypic diversity and horticultural taxonomy. In: Grumet R, Katzir N, Garcia-Mas J, eds., Genetics and Genomics of Cucurbitaceae. Springer, Cham. pp. 25–60.

Reddy A S, Marquez Y, Kalyna M, Barta A. 2013. Complexity of the alternative splicing landscape in plants. The Plant Cell, 25, 3657–3683.

Schmittgen T D, Livak K J. 2008. Analyzing real-time PCR data by the comparative CT method. Nature Protocols, 3, 1101–1108.

Shen J, Xu X, Zhang Y, Niu X, Shou W. 2021. Genetic Mapping and identification of the candidate genes for mottled rind in Cucumis melo L. Frontiers in Plant Science, 12, 769989.

Shi J, Yu H, Fu Y, Wang T, Zhang Y, Huang J, Li S, Zheng T, Ni X, Zhao J. 2023. Development and validation of functional kompetitive allele-specific PCR markers for herbicide resistance in Brassica napus. Frontiers in Plant Science, 14, 1213476.

Shin A Y, Koo N, Kim S, Sim Y M, Choi D, Kim Y M, Kwon S Y. 2019. Draft genome sequences of two oriental melons, Cucumis melo L. var. makuwa. Scientific Data, 6, 220.

Sun W, Li X, Huang H, Wei J, Zeng F, Huang Y, Sun Q, Miao W, Tian Y, Li Y, Gao L, Li X, Gao H. 2023. Mutation of CsARC6 affects fruit color and increases fruit nutrition in cucumber. Theoretical and Applied Genetics, 136, 111.

Tadmor Y, Burger J, Yaakov I. 2010. Genetics of flavonoid, carotenoid, and chlorophyll pigments in melon fruit rinds. Journal of Agricultural and Food Chemistry, 58, 10722–10728.

Tang H Y, Dong X, Wang J K, Xia J H, Xie F, Zhang Y, Yao X, Xu Y J, Wang Z J. 2018. Fine mapping and candidate gene prediction for white immature fruit skin in cucumber (Cucumis sativus L.). International Journal of Molecular Sciences, 19, 1493.

Tian S, Yang J, Fu Y, Zhang X, Zhang J, Zhao H, Hu Q, Liu P, He W, Han X, Wen C. 2023. McAPRR2: The key regulator of domesticated pericarp color in bitter gourd. Plants (Basel), 12, 3585.

Walker A R, Lee E, Bogs J, McDavid D A, Thomas M R, Robinson S P. 2007. White grapes arose through the mutation of two similar and adjacent regulatory genes. The Plant Journal, 49, 772–785.

Wang M, Wang P, Liang F, Ye Z, Li J, Shen C, Pei L, Wang F, Hu J, Tu L, Lindsey K, He D, Zhang X. 2018. A global survey of alternative splicing in allopolyploid cotton: Landscape, complexity and regulation. New Phytologist, 217, 163–178.

Xu X, Shen J, Zhang Y, Li G, Niu X, Shou W. 2021. Fine mapping of an immature rind color gene GR in melon. Scientia Agricultura Sinica, 54, 3308–3319. (in Chinese)

Zhan J, Zhong J, Cheng J, Wang Y, Hu K. 2023. Map-based cloning of the APRR2 gene controlling green stigma in bitter gourd (Momordica charantia). Frontiers in Plant Science, 14, 1128926.

Zhang H, Li X, Yu H, Zhang Y, Li M, Wang H, Wang D, Wang H, Fu Q, Liu M, Ji C, Ma L, Tang J, Li S, Miao J, Zheng H, Yi H. 2019. A high-quality melon genome assembly provides insights into genetic basis of fruit trait improvement. iScience, 22, 16–27.

Zhao G, Lian Q, Zhang Z, Fu Q, He Y, Ma S, Ruggieri V, Monforte A J, Wang P, Julca I, Wang H, Liu J, Xu Y, Wang R, Ji J, Xu Z, Kong W, Zhong Y, Shang J, Pereira L, et al. 2019. A comprehensive genome variation map of melon identifies multiple domestication events and loci influencing agronomic traits. Nature Genetics, 51, 1607–1615.

甜瓜果皮颜色基因的鉴定与标记开发

Identification and molecular marker development for peel color gene in melon (Cucumis melo L.)

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