黄芪种子萌发及萌发后生长过程中黄酮类化合物合成的动态变化
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摘要
在对膜荚黄芪和蒙古黄芪种子萌发和萌发后生长时期进行划分的基础上,研究了黄芪中3种主要黄酮类化合物芒柄花素、毛蕊异黄酮和毛蕊异黄酮苷在萌发和萌发后生长过程中含量的变化。为了弄清黄酮含量变化的机制,我们还对黄芪中黄酮生物合成途径酶的转录水平做了研究。结果表明,在种子萌发和萌发后生长阶段,两种黄芪中均出现了明显的黄酮类化合物累积增加的情况。通过检测黄酮类物质生物合成途径酶基因的转录表达水平,我们发现萌发阶段蒙古黄芪中黄酮类物质积累增加主要由上游途径酶基因的表达水平升高引起,而膜荚黄芪中则几乎涉及到所有途径酶。此外,我们还发现萌发后生长阶段中,两种黄芪中合成途径酶基因转录对黄酮类物质含量的调控则选择性地存在时间延迟效应。
Based on the stage definitions of seed germination and post-germination growth,we investigated the variations of contents of three major flavonoids during the two stages in Astragalus membranaceus and its variety of mongholicus. Besides,in order to understand how the variations were induced,the transcriptions of pathway enzyme involving in flavonoid biosynthesis were profiled. The contents of flavonoids increased during the two stages in both A. membranaceus and the variety. With the detection of gene transcriptions of the pathway enzymes,only the enzymes in upstream of the pathway contributed to the increase of flavonoids in the variety of mongholicus during the germination,whereas it was that almost all the pathway enzymes were involved in A.membranaceus. The transcriptions of pathway enzymes regulated the accumulations of flavonoids during postgermination growth,selectively with a pattern of time delay.
Based on the stage definitions of seed germination and post-germination growth,we investigated the variations of contents of three major flavonoids during the two stages in Astragalus membranaceus and its variety of mongholicus. Besides,in order to understand how the variations were induced,the transcriptions of pathway enzyme involving in flavonoid biosynthesis were profiled. The contents of flavonoids increased during the two stages in both A. membranaceus and the variety. With the detection of gene transcriptions of the pathway enzymes,only the enzymes in upstream of the pathway contributed to the increase of flavonoids in the variety of mongholicus during the germination,whereas it was that almost all the pathway enzymes were involved in A.membranaceus. The transcriptions of pathway enzymes regulated the accumulations of flavonoids during postgermination growth,selectively with a pattern of time delay.
引文
1.Yao M C,Qi Y,Bi K S,et al.A precolumn derivatization high-performance liquid chromatographic method with improved sensitivity and specificity for the determination of AstragalosideⅣin Radix astragali[J].Journal of Chromatographic Science,2000,38(8):325-328.
2.Lee D Y,Noh H J,Choi J,et al.Anti-inflammatory cycloartane-type saponins of Astragalus membranaceus[J].Molecules,2013,18(4):3725-3732.
3.Du H W,Zhao X L,Zhang A H,et al.Rapid separation,identification and analysis of Astragalus membranaceus Fisch using liquid chromatography-tandem mass spectrometry[J].Journal of Chromatographic Science,2014,52(3):226-231.
4.张冬青,汪德清.黄芪总黄酮生物学活性作用研究进展[J].中国中药杂志,2010,35(2):253-256.Zhang D Q,Wang D Q.Progressive studies on biological activity of total flavonoids of Astragalus[J].China Journal of Chinese Materia Medica,2010,35(2):253-256.
5.Zhang H X,Irving L J,Mcgill C,et al.The effects of salinity and osmotic stress on barley germination rate:sodium as an osmotic regulator[J].Annals of Botany,2010,106(6):1027-1035.
6.Ye N H,Zhu G H,Liu Y G,et al.Ascorbic acid and reactive oxygen species are involved in the inhibition of seed germination by abscisic acid in rice seeds[J].Journal of Experimental Botany,2012,63(5):1809-1822.
7.Ishibashi Y,Kasa S,Sakamoto M,et al.A role for reactive oxygen species produced by NADPH oxidases in the embryo and aleurone cells in barley seed germination[J].PLo S One,2015,10(11):e0143173.
8.Wojtyla,Lechowska K,Kubala S,et al.Different modes of hydrogen peroxide action during seed germination[J].Frontiers in Plant Science,2016,7:66.
9.Bailly C.Active oxygen species and antioxidants in seed biology[J].Seed Science Research,2004,14(2):93-107.
10.Zhang Y,Chen B X,Xu Z J,et al.Involvement of reactive oxygen species in endosperm cap weakening and embryo elongation growth during lettuce seed germination[J].Journal of Experimental Botany,2014,65(12):3189-3200.
11.Gapper C,Dolan L.Control of plant development by reactive oxygen species[J].Plant Physiology,2006,141(2):341-345.
12.Apel K,Hirt H.Reactive oxygen species:metabolism,oxidative stress,and signal transduction[J].Annual Review of Plant Biology,2004,55:373-399.
13.Kubasek W L,Shirley B W,Mckillop A,et al.Regulation of flavonoid biosynthetic genes in germinating Arabidopsis seedlings[J].The Plant Cell,1992,4(10):1229-1236.
14.Due1as M,Hernández T,Estrella I,et al.Germination as a process to increase the polyphenol content and antioxidant activity of lupin seeds(Lupinus angustifolius L.)[J].Food Chemistry,2009,117(4):599-607.
15.徐恒恒,黎妮,刘树君,等.种子萌发及其调控的研究进展[J].作物学报,2014,40(7):1141-1156.Xu H H,Li N,Liu S J,et al.Research progress in seed germination and its control[J].Acta Agronomica Sinica,2014,40(7):1141-1156.
16.段琦梅,梁宗锁,慕小倩,等.黄芪种子萌发特性的研究[J].西北植物学报,2005,25(6):1246-1249.Duan Q M,Liang Z S,Mu X Q,et al.Germination characteristics of Astragalus membranaceus seeds[J].Acta Botanica Boreali-Occidentalia Sinica,2005,25(6):1246-1249.
17.Kim Y B,Thwe A A,Li X H,et al.Accumulation of flavonoids and related gene expressions in different organs of Astragalus membranaceus Bge[J].Applied Biochemistry and Biotechnology,2014,173(8):2076-2085.
18.李景欣,高春宇,毕晓秀.Na Cl胁迫对黄芪种子萌发及幼苗生长的影响[J].内蒙古林业科技,2005,(3):11-14.Li J X,Gao C Y,Bi X X.Impaction to the Na Cl stress in the stage of seed germination and Seedlings of Radis astragali[J].Inner Mongolia Forestry Science&Technology,2005,(3):11-14.
19.Shibata T,Sakai E,Shimomura K.Effect of rapid freezing and thawing on hard-seed breaking in Astragalus mongholicus Bunge(Leguminosae)[J].Journal of Plant Physiology,1995,147(1):127-131.
20.郭凤霞,武志江,陈垣,等.蒙古黄芪种子水浸液对不同种党参种子萌发及幼苗生长的影响[J].中国中药杂志,2012,37(22):3375-3380.Guo F X,Wu Z J,Chen H,et al.Effect of Astragalus membranaceus var.mongholicus seed extracts on seed germination and seedling growth of different Codonopsis pilosula caltiver[J].China Journal of Chinese Materia Medica,2012,37(22):3375-3380.
21.张琳,秦雪梅.黄芪种子萌发实验[J].中药材,1994,17(11):9-10.Zhang L,Qin X M.Germination experiment of Astragalus membranaceus seeds[J].Journal of Chinese Medicinal Materials,1994,17(11):9-10.
22.Wang M,Van Der Meulen R M,Visser K,et al.Effects of dormancy-breaking chemicals on ABA levels in barley grain embryos[J].Seed Science Research,1998,8(2):129-137.
23.Liu X J,Xing D,Li L L,et al.Rapid determination of seed vigor based on the level of superoxide generation during early imbibitions[J].Photochemical&Photobiological Sciences,2007,6(7):767-774.
24.Oracz K,El-Maarouf-Bouteau H,Kranner I,et al.The mechanisms involved in seed dormancy alleviation by hydrogen cyanide unravel the role of reactive oxygen species as key factors of cellular signaling during germination[J].Plant Physiology,2009,150(1):494-505.
25.Müller K,Linkies A,Vreeburg R A M,et al.In vivo cell wall loosening by hydroxyl radicals during cress seed germination and elongation growth[J].Plant Physiology,2009,150(4):1855-1865.
26.El-Maarouf-Bouteau H,Bailly C.Oxidative signaling in seed germination and dormancy[J].Plant Signaling&Behavior,2008,3(3):175-182.
27.Tommasi F,Paciolla C,De Pinto M C,et al.A comparative study of glutathione and ascorbate metabolism during germination of Pinus pinea L.seeds[J].Journal of Experimental Botany,2001,52(361):1647-1654.
28.Diaz-Vivancos P,Barba-Espín G,Hernández J A.Elucidating hormonal/ROS networks during seed germination:insights and perspectives[J].Plant Cell Reports,2013,32(10):1491-1502.
29.Kreuzaler F,Ragg H,Fautz E,et al.UV-induction of Chalcone synthase mRNA in cell suspension cultures of Petroselinum hortense[J].Proceedings of the National A-cademy of Sciences of the United States of America,1983,80(9):2591-2593.
30.Ryder T B,Hedrick S A,Bell J N,et al.Organization and differential activation of a gene family encoding the plant defense enzyme chalcone synthase in Phaseolus vulgaris[J].Molecular and General Genetic MGG,1987,210(2):219-233.
31.谢灵玲,赵武玲,沈黎明.光照对大豆叶片苯丙氨酸裂解酶(PAL)基因表达及异黄酮合成的调节[J].植物学通报,2000,17(5):443-449.Xie L L,Zhao W L,Shen L M.Light regulation of the expression of PAL gene in soybean leaves and isoflavone synthesis[J].Chinese Bulletin of Botany,2000,17(5):443-449.
32.欧阳光察,薛应龙.植物苯丙烷类代谢的生理意义及其调控[J].植物生理学通讯,1988,24(3):9-16.Ouyang G C,Xue Y L.Physiological role and regulation of phenylpropanoid metabolism in plant[J].Plant Physiology Communications,1988,24(3):9-16.
33.Xu R Y,Nan P,Yang Y X,et al.Ultraviolet irradiation induces accumulation of isoflavonoids and transcription of genes of enzymes involved in the calycosin-7-O-β-D-glucoside pathway in Astragalus membranaceus Bge.var.mongholicus(Bge.)Hsiao[J].Physiologia Plantarum,2011,142(3):265-273.
34.李琳玲,程华,许锋,等.植物查尔酮异构酶研究进展[J].生物技术通讯,2008,19(6):935-937.Li L L,Cheng H,Xu F,et al.Progress of chalcone isomerase in plants[J].Letters in Biotechnology,2008,19(6):935-937.
2.Lee D Y,Noh H J,Choi J,et al.Anti-inflammatory cycloartane-type saponins of Astragalus membranaceus[J].Molecules,2013,18(4):3725-3732.
3.Du H W,Zhao X L,Zhang A H,et al.Rapid separation,identification and analysis of Astragalus membranaceus Fisch using liquid chromatography-tandem mass spectrometry[J].Journal of Chromatographic Science,2014,52(3):226-231.
4.张冬青,汪德清.黄芪总黄酮生物学活性作用研究进展[J].中国中药杂志,2010,35(2):253-256.Zhang D Q,Wang D Q.Progressive studies on biological activity of total flavonoids of Astragalus[J].China Journal of Chinese Materia Medica,2010,35(2):253-256.
5.Zhang H X,Irving L J,Mcgill C,et al.The effects of salinity and osmotic stress on barley germination rate:sodium as an osmotic regulator[J].Annals of Botany,2010,106(6):1027-1035.
6.Ye N H,Zhu G H,Liu Y G,et al.Ascorbic acid and reactive oxygen species are involved in the inhibition of seed germination by abscisic acid in rice seeds[J].Journal of Experimental Botany,2012,63(5):1809-1822.
7.Ishibashi Y,Kasa S,Sakamoto M,et al.A role for reactive oxygen species produced by NADPH oxidases in the embryo and aleurone cells in barley seed germination[J].PLo S One,2015,10(11):e0143173.
8.Wojtyla,Lechowska K,Kubala S,et al.Different modes of hydrogen peroxide action during seed germination[J].Frontiers in Plant Science,2016,7:66.
9.Bailly C.Active oxygen species and antioxidants in seed biology[J].Seed Science Research,2004,14(2):93-107.
10.Zhang Y,Chen B X,Xu Z J,et al.Involvement of reactive oxygen species in endosperm cap weakening and embryo elongation growth during lettuce seed germination[J].Journal of Experimental Botany,2014,65(12):3189-3200.
11.Gapper C,Dolan L.Control of plant development by reactive oxygen species[J].Plant Physiology,2006,141(2):341-345.
12.Apel K,Hirt H.Reactive oxygen species:metabolism,oxidative stress,and signal transduction[J].Annual Review of Plant Biology,2004,55:373-399.
13.Kubasek W L,Shirley B W,Mckillop A,et al.Regulation of flavonoid biosynthetic genes in germinating Arabidopsis seedlings[J].The Plant Cell,1992,4(10):1229-1236.
14.Due1as M,Hernández T,Estrella I,et al.Germination as a process to increase the polyphenol content and antioxidant activity of lupin seeds(Lupinus angustifolius L.)[J].Food Chemistry,2009,117(4):599-607.
15.徐恒恒,黎妮,刘树君,等.种子萌发及其调控的研究进展[J].作物学报,2014,40(7):1141-1156.Xu H H,Li N,Liu S J,et al.Research progress in seed germination and its control[J].Acta Agronomica Sinica,2014,40(7):1141-1156.
16.段琦梅,梁宗锁,慕小倩,等.黄芪种子萌发特性的研究[J].西北植物学报,2005,25(6):1246-1249.Duan Q M,Liang Z S,Mu X Q,et al.Germination characteristics of Astragalus membranaceus seeds[J].Acta Botanica Boreali-Occidentalia Sinica,2005,25(6):1246-1249.
17.Kim Y B,Thwe A A,Li X H,et al.Accumulation of flavonoids and related gene expressions in different organs of Astragalus membranaceus Bge[J].Applied Biochemistry and Biotechnology,2014,173(8):2076-2085.
18.李景欣,高春宇,毕晓秀.Na Cl胁迫对黄芪种子萌发及幼苗生长的影响[J].内蒙古林业科技,2005,(3):11-14.Li J X,Gao C Y,Bi X X.Impaction to the Na Cl stress in the stage of seed germination and Seedlings of Radis astragali[J].Inner Mongolia Forestry Science&Technology,2005,(3):11-14.
19.Shibata T,Sakai E,Shimomura K.Effect of rapid freezing and thawing on hard-seed breaking in Astragalus mongholicus Bunge(Leguminosae)[J].Journal of Plant Physiology,1995,147(1):127-131.
20.郭凤霞,武志江,陈垣,等.蒙古黄芪种子水浸液对不同种党参种子萌发及幼苗生长的影响[J].中国中药杂志,2012,37(22):3375-3380.Guo F X,Wu Z J,Chen H,et al.Effect of Astragalus membranaceus var.mongholicus seed extracts on seed germination and seedling growth of different Codonopsis pilosula caltiver[J].China Journal of Chinese Materia Medica,2012,37(22):3375-3380.
21.张琳,秦雪梅.黄芪种子萌发实验[J].中药材,1994,17(11):9-10.Zhang L,Qin X M.Germination experiment of Astragalus membranaceus seeds[J].Journal of Chinese Medicinal Materials,1994,17(11):9-10.
22.Wang M,Van Der Meulen R M,Visser K,et al.Effects of dormancy-breaking chemicals on ABA levels in barley grain embryos[J].Seed Science Research,1998,8(2):129-137.
23.Liu X J,Xing D,Li L L,et al.Rapid determination of seed vigor based on the level of superoxide generation during early imbibitions[J].Photochemical&Photobiological Sciences,2007,6(7):767-774.
24.Oracz K,El-Maarouf-Bouteau H,Kranner I,et al.The mechanisms involved in seed dormancy alleviation by hydrogen cyanide unravel the role of reactive oxygen species as key factors of cellular signaling during germination[J].Plant Physiology,2009,150(1):494-505.
25.Müller K,Linkies A,Vreeburg R A M,et al.In vivo cell wall loosening by hydroxyl radicals during cress seed germination and elongation growth[J].Plant Physiology,2009,150(4):1855-1865.
26.El-Maarouf-Bouteau H,Bailly C.Oxidative signaling in seed germination and dormancy[J].Plant Signaling&Behavior,2008,3(3):175-182.
27.Tommasi F,Paciolla C,De Pinto M C,et al.A comparative study of glutathione and ascorbate metabolism during germination of Pinus pinea L.seeds[J].Journal of Experimental Botany,2001,52(361):1647-1654.
28.Diaz-Vivancos P,Barba-Espín G,Hernández J A.Elucidating hormonal/ROS networks during seed germination:insights and perspectives[J].Plant Cell Reports,2013,32(10):1491-1502.
29.Kreuzaler F,Ragg H,Fautz E,et al.UV-induction of Chalcone synthase mRNA in cell suspension cultures of Petroselinum hortense[J].Proceedings of the National A-cademy of Sciences of the United States of America,1983,80(9):2591-2593.
30.Ryder T B,Hedrick S A,Bell J N,et al.Organization and differential activation of a gene family encoding the plant defense enzyme chalcone synthase in Phaseolus vulgaris[J].Molecular and General Genetic MGG,1987,210(2):219-233.
31.谢灵玲,赵武玲,沈黎明.光照对大豆叶片苯丙氨酸裂解酶(PAL)基因表达及异黄酮合成的调节[J].植物学通报,2000,17(5):443-449.Xie L L,Zhao W L,Shen L M.Light regulation of the expression of PAL gene in soybean leaves and isoflavone synthesis[J].Chinese Bulletin of Botany,2000,17(5):443-449.
32.欧阳光察,薛应龙.植物苯丙烷类代谢的生理意义及其调控[J].植物生理学通讯,1988,24(3):9-16.Ouyang G C,Xue Y L.Physiological role and regulation of phenylpropanoid metabolism in plant[J].Plant Physiology Communications,1988,24(3):9-16.
33.Xu R Y,Nan P,Yang Y X,et al.Ultraviolet irradiation induces accumulation of isoflavonoids and transcription of genes of enzymes involved in the calycosin-7-O-β-D-glucoside pathway in Astragalus membranaceus Bge.var.mongholicus(Bge.)Hsiao[J].Physiologia Plantarum,2011,142(3):265-273.
34.李琳玲,程华,许锋,等.植物查尔酮异构酶研究进展[J].生物技术通讯,2008,19(6):935-937.Li L L,Cheng H,Xu F,et al.Progress of chalcone isomerase in plants[J].Letters in Biotechnology,2008,19(6):935-937.