Analysis of the upstream region of the MYB10 gene in the anthocyanin biosynthesis pathway of Iranian red-fleshed apples using polymerase chain reaction (PCR)

Document Type : Original Article

Authors

1 Associate Professor, Department of Horticultural Sciences, Gorgan University of Agricultural Sciences and Natural Resources, Gorgan, Iran.

2 M.Sc. Graduate in Pomology, Department of Horticultural Sciences, Gorgan University of Agricultural Sciences and Natural Resources, Gorgan, Iran.

3 Assistant Professor, Department of Plant Breeding and Biotechnology, Gorgan University of Agricultural Sciences and Natural Resources, Gorgan, Iran.

10.30466/rip.2023.54076.1226

Abstract

Iran is an important center of genetic diversity in apple, and red-fleshed apples occur in several Iranian regions, including Taleghan, Mazandaran, and Shahroud. This study aimed to identify single- and double-nucleotide differences in the upstream region of the MYB10 gene, which encodes a transcription factor involved in anthocyanin biosynthesis. A portion of the gene's upstream region was amplified and sequenced using a specific polymerase chain reaction (PCR) primer. PCR products were separated by agarose gel electrophoresis, and the sizes of the amplified bands in red-fleshed apple genotypes were compared with the known allele sizes of reference foreign apple cultivars and a base-pair marker. Based on sequence variations in the upstream region of MYB10, the Mazand 2, SH10, and GH2 apple genotypes were assigned to the same group. Although these genotypes differed in point mutations within the upstream region, red-fleshed apples exhibited greater similarity to one another than to white-fleshed apples. These results indicate a distinct organization of the upstream region of this gene in red-fleshed apples and suggest that native Iranian red-fleshed cultivars differ from the red-fleshed apple cultivar originating from Kazakhstan. This finding points to the potential existence of novel red-fleshed apple genotypes. Overall, the study revealed a high level of polymorphism among apple cultivars and Iranian red-fleshed apple genotypes, supporting the view that Iran may be one of the world's centers of apple diversity. These findings may contribute to the introduction of native Iranian red-fleshed apple cultivars to consumer markets through breeding programs.

Keywords

References

فرامرزی، ش. 1389. مطالعه صفات مرفولوژیک و تنوع ژنتیکی سیب­های گوشت قرمز ایرانی با استفاده از نشانگرهای ریزماهواره. پایان نامه کارشناسی­ارشد دانشگاه تربیت مدرس.
فرامرزی، ش.، یدالهی، ع.، برزگر، م. و کریم­زاده، ق. 1394. مقایسه مهم‌ترین ویژگی‌های فیزیکوشیمیایی میوه سیب گوشت قرمز و تعدادی از ارقام سیب گلاب. به‌زراعی کشاورزی، 17(3): 729-741.
فرامرزی، ش.، یدالهی، ع.، حاج­نجاری، ح.، شجاعیان، ع. و دامیار، س. 1393. بررسی صفات مورفولوژیک سیب‌های گوشت قرمز ایرانی در مقایسه با تعدادی از ارقام بومی و تجاری. به‌زراعی کشاورزی، 16(1): 1-10.
میرمحمدی­میبدی، ع،م. 1382. اصلاح نباتات در باغبانی. انتشارات دانشگاه صنعتی اصفهان، 224 ص.
Allan, A.C., Hellens, R.P. and Laing, W.A., 2008. MYB transcription factors that colour our fruit. Trends in plant science, 13(3): 99-102.
Andersen, O.M. and Jordhein, M. 2006. The anthocyanins. In: O.M. Andersen and K.R. Markham (Eds.). Flavonoids: Chemistry, Biochemistry and Applications. London, CRC Press. 471-552.
Ban, Y., Honda, C., Hatsuyama, Y., Igarashi, M., Bessho, H. and Moriguchi, T. 2007. Isolation and functional analysis of a MYB transcription factor gene that is a key regulator for the development of red coloration in apple skin. Plant and cell physiology, 48(7): 958-970.
Bhandari, H.R., Bhanu, A.N., Srivastava, K., Singh, M.N. and Shreya, H.A. 2017. Assessment of genetic diversity in crop plants-an overview. Advances in Plants and Agriculture Research, 7(3): 279-286.
Bouvier, L., Lespinasse, Y. and Schuster, M., 1999, September. Karyotype analysis of a haploid plant of apple (Malus domestica). In Eucarpia symposium on Fruit Breeding and Genetics 538 (pp. 321-324).
Breseghello, F. and Sorrells, M.E., 2006. Association analysis as a strategy for improvement of quantitative traits in plants. Crop Science, 46(3): 1323-1330.
Bu, Y., Wu, X., Sun, N., Man, Y. and Jing, Y. 2021. Codon usage bias predicts the functional MYB10 gene in Populus. Journal of plant physiology, 265: 153491.
Chagné, D., Carlisle, C.M., Blond, C., Volz, R.K., Whitworth, C.J., Oraguzie, N.C., Crowhurst, R.N., Allan, A.C., Espley, R.V., Hellens, R.P. anگاه تربیت مدرسd Gardiner, S.E. 2007. Mapping a candidate gene (MdMYB10) for red flesh and foliage colour in apple. BMC genomics, 8(212): 1-11.
Channuntapipat, C., Sedgley, M. and Collins, G. 2001. Sequences of the cDNAs and genomic DNAs encoding the S1, S7, S8, and Sf alleles from almond, Prunus dulcis. Theoretical and Applied Genetics, 103: 1115-1122.
Chen, L., Hu, B., Qin, Y., Hu, G. and Zhao, J. 2019. Advance of the negative regulation of anthocyanin biosynthesis by MYB transcription factors. Plant physiology and biochemistry, 136: 178-187.
D’Abrosca, B., Pacifico, S., Cefarelli, G., Mastellone, C. and Fiorentino, A. 2007. ‘Limoncella’ apple, an Italian apple cultivar: Phenolic and flavonoid contents and antioxidant activity. Food chemistry, 104(4): 1333-1337.
Doyle, J.J. and Doyle, J.L., 1987. A rapid DNA isolation procedure for small quantities of fresh leaf tissue. Phytochemical bulletin.
Espley, R.V., Brendolise, C., Chagne, D., Kutty-Amma, S., Green, S., Volz, R., Putterill, J., Schouten, H.J., Gardiner, S.E., Hellens, R.P. and Allan, A.C. 2009. Multiple repeats of a promoter segment causes transcription factor autoregulation in red apples. The Plant Cell, 21(1): 168-183.
Espley, R.V., Hellens, R.P., Putterill, J., Stevenson, D.E., Kutty‐Amma, S. and Allan, A.C. 2007. Red colouration in apple fruit is due to the activity of the MYB transcription factor, MdMYB10. The Plant Journal, 49(3): 414-427.
Fang, Z.Z., Lin-Wang, K., Zhou, D.R., Lin, Y.J., Jiang, C.C., Pan, S.L., Espley, R.V., Andre, C.M. and Ye, X.F. 2021. Activation of PsMYB10. 2 transcription causes anthocyanin accumulation in flesh of the red-fleshed mutant of ‘Sanyueli’ (Prunus salicina Lindl.). Frontiers in Plant Science, 12: p.680469.
Gantet, P. and Memelink, J. 2002. Transcription factors: tools to engineer the production of pharmacologically active plant metabolites. Trends in Pharmacological Sciences, 23(12): 563-569.
Govindaraj, M., Vetriventhan, M. and Srinivasan, M. 2015. Importance of genetic diversity assessment in crop plants and its recent advances: an overview of its analytical perspectives. Genetics research international, 2015. 1-14.
Harris, S.A., Robinson, J.P. and Juniper, B.E., 2002. Genetic clues to the origin of the apple. TRENDS in Genetics, 18(8): 426-430.
Hokanson, S.C., Szewc-McFadden, A.K., Lamboy, W.F. and McFerson, J.R. 1998. Microsatellite (SSR) markers reveal genetic identities, genetic diversity and relationships in a Malus× domestica Borkh. core subset collection. Theoretical and applied genetics, 97: 671-683.
Janick, J., Cummins, J.N., Susan, K., Brown, S.K. and Hemmat, M. 1996. Apple In: Fruit Breeding, Vol. 1: تنوTree and Tropical Fruit, J. Janick and Moore, J.N. (eds.) John Wiley and Sons, Inc. 1-77.
Jiang, S., Chen, M., He, N., Chen, X., Wang, N., Sun, Q., Zhang, T., Xu, H., Fang, H., Wang, Y. and Zhang, Z. 2019. MdGSTF6, activated by MdMYB1, plays an essential role in anthocyanin accumulation in apple. Horticulture Research, 6, p.40.
Joshi, S.P., Ranjekar, P.K. and Gupta, V.S. 1999. Molecular markers in plant genome analysis. Current Science, 77: 230-240.
Kando, S. 2009. Fruit ripening and characteristics regulated by physiologically active substances. Fresh Produce Journal (Special Issue), 17: 7-11.
Koes, R.E., Quattrocchio, F. and Mol, J.N., 1994. The flavonoid biosynthetic pathway in plants: function and evolution. BioEssays, 16(2), pp.123-132.
Kong, J.M., Chia, L.S., Goh, N.K., Chia, T.F. and Brouillard, R., 2003. Analysis and biological activities of anthocyanins. Phytochemistry, 64(5): 923-933.
Lancaster, J.E. and Dougall, D.K., 1992. Regulation of skin color in apples. Critical Reviews in Plant Sciences, 10(6), pp.487-502.
Lespinasse, Y., Bouvier, L., Djulbic, M. and Chevreau, E., 1996. Haploidy in apple and pear. In Eucarpia Symposium on Fruit Breeding and Genetics 484: 49-54.
Lin-Wang, K., Bolitho, K., Grafton, K., Kortstee, A., Karunairetnam, S., McGhie, T.K., Espley, R.V., Hellens, R.P. and Allan, A.C. 2010. An R2R3 MYB transcription factor associated with regulation of the anthocyanin biosynthetic pathway in Rosaceae. BMC plant biology, 10: 1-17.
Mahmoudi, E., Mohammad, S.B., Yadollahi, A. and Hosseini, E. 2012. Independence of color intensity variation in red flesh apples from the number of repeat units in promoter region of the MdMYB10 gene as an allele to MdMYB1 and MdMYBA. Iranian Journal of Biotechnology, 10: 153-160.
Martin, C. and Paz-Ares, J., 1997. MYB transcription factors in plants. Trends in Genetics, 13(2), pp.67-73.
Medina-Puche, L., Cumplido-Laso, G., Amil-Ruiz, F., Hoffmann, T., Ring, L., Rodríguez-Franco, A., Caballero, J.L., Schwab, W., Muñoz-Blanco, J. and Blanco-Portales, R. 2014. MYB10 plays a major role in the regulation of flavonoid/phenylpropanoid metabolism during ripening of Fragaria×ananassa fruits. Journal of experimental botany, 65(2): 401-417.
Plunkett, B.J., Henry-Kirk, R., Friend, A., Diack, R., Helbig, S., Mouhu, K., Tomes, S., Dare, A.P., Espley, R.V., Putterill, J. and Allan, A.C. 2019. Apple B-box factors regulate light-responsive anthocyanin biosynthesis genes. Scientific reports, 9(1):17762.
Raddova, J., Beránek, M., Oukropec, I. and Vachůn, M. 2003. RAPD analysis of peaches within Czech National Collection. Czech Journal of Genetics and Plant Breeding, 39(4): 113.
Ramsay, N.A. and Glover, B.J. 2005. MYB–bHLH–WD40 protein complex and the evolution of cellular diversity. Trends in plant science, 10(2): 63-70.
Riechmann, J.L, Heard, J.,   Martin, G Reuber, L.C.-Z. Jiang , C-Z Keddie,J Adam,L., Pineda,O  Ratcliffe,O.J and Yu, G.L2000. Arabidopsis Transcription Factors: Genome-Wide Comparative Analysis Among Eukaryotes.Science 5490 (299):2105-2110.
Romero, I., Fuertes, A., Benito, M.J., Malpica, J.M., Leyva, A., and Paz-Ares, J. 1998.  More than 80 R2R3-MYB regulatory genes in the genome of Arabidopsis thaliana. Plant Journal, 14(3): 273-284.
Rupasinghe, H.P., Huber, G.M., Embree, C. and Forsline, P.L 2010. Red-fleshed apple as a source for functional beverages. Canadian Journal of Plant Sciences, 90: 95-100.
Schwarz, H.P., Braun, P. and Keiciier, R. 2009. Red apple juice: breeding, drink-and growing technology for the development of a new, innovative product. Bulletin of the University of Agricultural Sciences & Veterinary Medicine Cluj-Napoca. Horticulture, 66(1): 218-231.
Stracke, R., Werber, M. and Weisshaar, B. 2001. The R2R3-MYB gene family in Arabidopsis thaliana. Current opinion in plant biology, 4(5): 447-456.
Sun-Waterhouse, D., Luberriaga, C., Jin, D., Wibisono, R., Wadhwa, S.S. and Waterhouse, G.I.N. 2013. Juices, fibres and skin waste extracts from white, pink or red-fleshed apple genotypes as potential food ingredients: A comparative study. Food and Bioprocess Technology, 6: 377-390.
Takos, A.M., Jaffé, F.W., Jacob, S.R., Bogs, J., Robinson, S.P. and Walker, A.R. 2006. Light-induced expression of a MYB gene regulates anthocyanin biosynthesis in red apples. Plant physiology, 142(3) 1216-1232.
Tatum, T.C., Stepanovic, S., Biradar, D.P., Rayburn, A.L. and Korban, S.S. 2005. Variation in nuclear DNA content in Malus species and cultivated apples. Genome, 48(5): 924-930.
Tsao, R., Yang, R., Young, J.C. and Zhu, H., 2003. Polyphenolic profiles in eight apple cultivars using high-performance liquid chromatography (HPLC). Journal of agricultural and food chemistry, 51(21): 6347-6353.
Tzulker, R., Glazer, I., Bar-Ilan, I., Holland, D., Aviram, M. and Amir, R. 2007. Antioxidant activity, polyphenol content, and related compounds in different fruit juices and homogenates prepared from 29 different pomegranate accessions. Journal of agricultural and food chemistry, 55(23): 9559-9570.
Umikalsom, Y., Grayer-Barkmeijer, R.J. and Harborne, J.B., 1994. A comparison of the flavonoids in Athyriaceae and Aspleniaceae. Biochemical systematics and ecology, 22(6), pp.587-594.
Walker, A.R., Lee, E., Bogs, J., McDavid, D.A., Thomas, M.R. and Robinson, S.P. 2007. White grapes arose through the mutation of two similar and adjacent regulatory genes. The Plant Journal, 49(5) 772-785.
Wang, H., Zhang, H., Yang, Y., Li, M., Zhang, Y., Liu, J., Dong, J., Li, J., Butelli, E., Xue, Z. and Wang, A. 2020. The control of red colour by a family of MYB transcription factors in octoploid strawberry (Fragaria× ananassa) fruits. Plant Biotechnology Journal, 18(5) 1169-1184.
Wang, X.Q., Han, J., Wen, Y., Jiang, W.B., Fang, J.G., Zhang, B.B. and Ma, R.J. 2016. Cloning and bioinformatic analysis of transcription factor MYB10 from the red-leaf peach. Genetics and Molecular Research: GMR, 15(4).
Way, R.D., Aldwinckle, H.S., Lamb, R.C., Rejman, A., Sansavini, S., Shen, T., Watkins, R., Westwood, M.N. and Yoshida, Y. 1991. Apples (malus). Genetic Resources of Temperate Fruit and Nut Crops 290: 3-46.
Xi, W., Feng, J., Liu, Y., Zhang, S. and Zhao, G. 2019. The R2R3-MYB transcription factor PaMYB10 is involved in anthocyanin biosynthesis in apricots and determines red blushed skin. BMC plant biology, 19: 1-14.
Yan, H., Pei, X., Zhang, H., Li, X., Zhang, X., Zhao, M., Chiang, V.L., Sederoff, R.R. and Zhao, X. 2021. MYB-mediated regulation of anthocyanin biosynthesis. International Journal of Molecular Sciences, 22(6): 3103-3121.
Yanhui, C., Xiaoyuan, Y., Kun, H., Meihua, L., Jigang, L., Zhaofeng, G., Zhiqiang, L., Yunfei, Z., Xiaoxiao, W., Xiaoming, Q. and Yunping, S. 2006. The MYB transcription factor superfamily of Arabidopsis: expression analysis and phylogenetic comparison with the rice MYB family. Plant molecular biology, 60: 107-124.
Zhou-Zhi, Q. 1999. The apple genetic resources in China: The apple species and their distributions, informative characteristics and utilization. Genetic Resources and Crop Evolution, 46: 599-609.
Research in Pomology
Volume 8, Issue 2
March 2024
Pages 44-58

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