Evaluating Expression of Gene Encoding Flavonol Synthase in Green TrungDu and Purple TrungDu by Real time PCR and HPLC Technique
Main Article Content
Abstract
In plants, flavonol synthase (FLS) is a multifunctional enzyme that converts dihydroflavonol into flavonols and naringenin into dihydrokaempferol. FLS from tea has been shown to metabolize dihydroquercetin to quercetin. In this study, we conducted studying on the relationship between quercetin content and the expression level of FLS in two traditional tea cultivars of Vietnam, TrungDuxanh and TrungDutim tea grown in tea garden of Thai Nguyen University of Agriculture and Forestry. Tea shoots with one apical bud and two to three young leaves using as research materials which collected in September 2017. The using of HPLC technique did not detect the quercetin content from the these tea samples. The preliminary results of quantification of FLS gene expression by real time PCR showed the expression level of FLS in green Trung Du tea is higher than purple Trung Du, although the difference is not great. Thus, at the time of collecting, the expression of FLS in green Trung Du and purple Trung Du can not synthesize quercetin but synthesize other flavonols.
Keywords:
TrungDu tea, green TrungDu, purple TrungDu, flavonol, quercetin, flavonol synthase.
References
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[3] A.X. Cheng, X.J. Han, Y.F. Wu, H.X. Lou, The function and catalysis of 2-oxoglutarate-dependent oxygenases involved in plant flavonoid biosynthesis, Int J Mol Sci. 15 (2014) 1080-1095. https://doi.org/10.3390/ijms15011080.
[4] M.M. Berger, Can oxidative damage be treated nutritionally?, Clinical Nutrition, 24 (2005) 172-183. https://doi.org/10.1016/j.clnu.2004.10.003.
[5] J.A. Vita, Polyphenols and cardiovascular disease: effects on endothelial and platelet function, The American Journal of Clinical Nutrition 81 (2005) 292-297. https://doi.org/10.1093/ajcn/81.1.292S.
[6] B.H. Havsteen, The biochemistry and medical significance of the flavonoids, Pharmacol Ther 96 (2002) 67-202. https://doi.org/10.1016/s0163-7258(02)00298-x.
[7] E. Butelli, L. Titta, M. Giorgio, H.P. Mock, A. Matros, S. Peterek, E.G. Schijlen, R.D. Hall, A.G. Bovy, J. Luo, C. Martin, Enrichment of tomato fruit with health-promoting anthocyanins by expression of select transcription factors, Nat Biotechnol 26 (2008) 1301-1308. https://doi.org/ 10.1038/nbt.1506.
[8] S. Czemmel, R. Stracke, B. Weisshaar, N. Cordon, N.N. Harris, A.R. Walker, S.P. Robinson, J. Bogs, The grapevine R2R3-MYB transcription factor VvMYBF1 regulates flavonol synthesis in developing grape berries, Plant Physiol, 151 (2009) 1513-1530. https://doi.org/10.1104/pp.109. 142059
[9] X. He, X. Zhao, L. Gao, X. Shi, X. Dai, Y. Liu, T. Xia, Y. Wang, Isolation and Characterization of Key Genes that Promote Flavonoid Accumulation in Purple-leaf Tea (Camellia sinensis L.), Sci Rep 8 (2018) 130. https://doi.org/10.1038/s41598-017-18133-z.
[10] Y.B. Kim, K. Kim, Y. Kim, P.A. Tuan, H.H. Kim, J.W. Cho, S.U. Park, Cloning and characterization of a flavonol synthase gene from Scutellaria baicalensis, Scientific World Journal 2014 (2014) 980740. https://doi.org/10.1155/2014/980740.
[11] M.E. Harbowy, D.A. Balentine, Tea chemistry, Critical Reviews ill Plant Sciences, 16 (1997) 415-480. https://doi.org/10.1080/07352689709701956.
[12] D.L. Mckay, J.B. Blumberg, The role of tea in human health: an update, J Am Coll Nutr, 21 (2002) 1-13. https://doi.org/10.1080/07315724. 2002.10719187.
[13] T. Tohge, L.P. De Souza, A.R. Fernie, Current understanding of the pathways of flavonoid biosynthesis in model and crop plants, J Exp Bot 68 (2017) 4013-4028. https://doi.org/10.1093/ jxb/erx177.
[14] B. Winkel-Shirley, Flavonoid biosynthesis. A colorful model for genetics, biochemistry, cell biology, and biotechnology, Plant Physiol 126 (2001) 485-493. https://doi.org/10.1104/pp.126.2. 485.
[15] L. Wang, D. Pan, M. Liang, Y.S. Abubakar, J. Li, J. Lin, S. Chen, W. Chen, Regulation of Anthocyanin Biosynthesis in Purple Leaves of Zijuan Tea (Camellia sinensis var. kitamura), Int J Mol Sci 18 (2017) 833. https:// doi.org/10.3390/ ijms18040833.
[16] G.Z. Lin, Y.J. Lian, J.H. Ryu, M.K. Sung, J.S. Park, H.J. Park, B.K. Park, J.S. Shin, M.S. Lee, C.I. Cheon, Expression and purification of His-tagged flavonol synthase of Camellia sinensis from Escherichia coli, Protein Expr Purif 55 (2007) 287-292. https://doi.org/10.1016/j.pep.2007. 05.013.
[17] A. Finger, S. Kuhr, U.H. Engelhardt, Chromatography of tea constituents, J Chromatogr 624 (1992) 293-315. https://doi.org/10. 1016/0021-9673(92)85685-M.
[18] L.Z. Lin, P. Chen, J.M. Harnly, New phenolic components and chromatographic profiles of green and fermented teas, J Agric Food Chem. 56 (2008) 8130-8140. https://doi.org/10.1021/jf800 986s.
[19] X. Hao, D.P. Horvath, W.S. Chao, Y. Yang, X. Wang, B. Xiao, Identification and evaluation of reliable reference genes for quantitative real-time PCR analysis in tea plant (Camellia sinensis (L.) O. Kuntze), Int J Mol Sci. 15 (2014) 22155-22172. https://doi.org/10.3390/ijms151222155.
[20] M. Sun, Y. Wang, D. Yang, C. Wei, L. Gao, T. Xia, Y. Shan, Y. Luo, Reference genes for real-time fluorescence quantitative PCR in Camellia sinensis, Chinese Bulletin of Botany 45 (2010) 579-587. https://doi.org/10.3969/j.issn.1674-3466. 2010.05.007.
[21] Z.J. Wu, C. Tian, Q. Jiang, X.H. Li, J. Zhuang, Selection of suitable reference genes for qRT-PCR normalization during leaf development and hormonal stimuli in tea plant (Camellia sinensis), Sci Rep. 6 (2016) 19748. https://doi.org/10.1038/ srep19748.
[22] Y.S. Wang, L.P. Gao, Y. Shan, Y.J. Liu, Y.W. Tian, T. Xia, Influence of shade on flavonoid biosynthesis in tea (Camellia sinensis (L.) O. Kuntze), Scientia Horticulturae 141 (2012) 7–16. https://doi.org/10.1016/j.scienta.2012.04.013.
[23] M. Liu, H.L. Tian, J.H. Wu, R.R. Cang, R.X. Wang, X.H. Qi, Q. Xu, X.H. Chen, Relationship between gene expression and the accumulation of catechin during spring and autumn in tea plants (Camellia sinensis L.), Hortic Res. 2 (2015) 15011-15019. https://doi.org/10.1038/hortres.2015. 23.
[24] K. Wei, L. Wang, C. Zhang, L. Wu, H. Li, F. Zhang, H. Cheng, Transcriptome Analysis Reveals Key Flavonoid 3'-Hydroxylase and Flavonoid 3',5'-Hydroxylase Genes in Affecting the Ratio of Dihydroxylated to Trihydroxylated Catechins in Camellia sinensis, PLoS One 10 (2015) 137925-137937. https://doi.org/10.1371/ journal.pone.0137925.