大豆种子老化MDA和4-HNE的含量变化相关性研究
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摘要
研究了大豆"中豆27"种子在人工老化和自然老化过程中,电导率、丙二醛和4-羟基壬烯醛(4-hydroxy-(E)-2-nonenal,4-HNE)含量,4-HNE的组织特异性积累及其与合成清除酶系统相关的基因表达调控差异,以探寻大豆种子的老化生理指标。结果表明,伴随着老化,大豆种子萌发率逐渐降低,浸泡液电导率逐渐增大;丙二醛含量升高;4-羟基壬烯醛(4-hydroxy-(E)-2-nonenal,4-HNE)在低活力干种子(萌发率10%)中积累量显著升高,且在人工老化和自然老化至中等活力干种子(萌发率50%～80%)和吸胀20 h的种子中,前者含量皆高于后者;免疫组化染色结果表明,老化至萌发率80%的种子中的4-HNE积累量则是自然老化种子明显高于人工老化种子。同时,老化诱导4-HNE的合成,但与4-HNE清除相关的乙醇脱氢酶(alcohol dehydrogenase)和谷胱甘肽-S-转移酶(glutathione-S-transferase)基因在人工老化下表达水平显著高于自然老化,即2种老化条件下种子中4-HNE的清除可能存在差异。因此,丙二醛和4-HNE累积与种子活力呈负相关关系,可作为种子老化程度的生理指标。
To explore the physiological indicators reflecting soybean seed aging,in this paper,soybean variety "Zhongdou 27" was used as materials to measure the changes of electrical conductivity of seed soak solution, the content of MDA(malondialdehyde) and 4-HNE(4-hydroxy-(E)-2-nonenal),tissue specific accumulation of 4-HNE,and the differences in the expression and regulation of genes relative to 4-HNE formation and scavenging enzyme system under natural aging and artificial aging.The results showed that the germination rate of seed soak solution was decreased gradually,While the electrical conductivity and Malondialdehyde(MDA) content were increased.The accumulation of 4-hydroxy-(E)-2-nonenal(4-HNE) was increased significantly in low-activity dry seeds(germination rate 10%) during the artificial and natural aging process.The content of artificial and natural aging to medium vigor dry seeds(germination rate 50%—80%) and imbibition seeds for 20 h were higher than that of the latter.The content of 4-hydroxy-(E)-2-nonenal Immunohistochemical staining showed that the accumulation of 4-HNE in the seeds aged to 80% germination rate were significantly higher than that of the artificially aged seeds.At the same time,aging induces the synthesis of 4-HNE,but genes of alcohol dehydrogenase and glutathione-S transferase associating with scavenging 4-HNE expressed significantly higher in the artificially aged seeds than that of naturally aged seeds.That meant the scavenging of 4-HNE in seeds under the two aging conditions may be different.Therefore,MDA and 4-HNE accumulation were negatively correlated with seed vigor.It could be used as physiological indicators of seed aging.
To explore the physiological indicators reflecting soybean seed aging,in this paper,soybean variety "Zhongdou 27" was used as materials to measure the changes of electrical conductivity of seed soak solution, the content of MDA(malondialdehyde) and 4-HNE(4-hydroxy-(E)-2-nonenal),tissue specific accumulation of 4-HNE,and the differences in the expression and regulation of genes relative to 4-HNE formation and scavenging enzyme system under natural aging and artificial aging.The results showed that the germination rate of seed soak solution was decreased gradually,While the electrical conductivity and Malondialdehyde(MDA) content were increased.The accumulation of 4-hydroxy-(E)-2-nonenal(4-HNE) was increased significantly in low-activity dry seeds(germination rate 10%) during the artificial and natural aging process.The content of artificial and natural aging to medium vigor dry seeds(germination rate 50%—80%) and imbibition seeds for 20 h were higher than that of the latter.The content of 4-hydroxy-(E)-2-nonenal Immunohistochemical staining showed that the accumulation of 4-HNE in the seeds aged to 80% germination rate were significantly higher than that of the artificially aged seeds.At the same time,aging induces the synthesis of 4-HNE,but genes of alcohol dehydrogenase and glutathione-S transferase associating with scavenging 4-HNE expressed significantly higher in the artificially aged seeds than that of naturally aged seeds.That meant the scavenging of 4-HNE in seeds under the two aging conditions may be different.Therefore,MDA and 4-HNE accumulation were negatively correlated with seed vigor.It could be used as physiological indicators of seed aging.
引文
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[21]高华伟,满强,武猛,等.耐储藏大豆种质资源的筛选[J].大豆科学,2014,33(1):000006-000012.
[22]Bale2evic'Tubic'S,Tatic'M,or?evic'V,et al.Seed viability of oil crops depending on storage conditions[J].Helia,2010,33(52):153-160.
[23]周小梅,王自霞,乔燕祥.人工老化处理对芝麻种子生理生化特性的影响[J].中国油料作物学报,2008,30(4):460-463.
[24]张瑛,滕斌,吴敬德,等.水稻种子高温高湿人工加速老化试验方法研究[J].中国粮油学报,2010,25(10):8-12.
[25]李稳香,颜启传.杂交水稻自然老化种子与人工老化种子性能差异研究[J].杂交水稻,1997(3):26-28.
[2]徐是雄,唐锡华,傅家瑞.种子生理的研究进展[M].广州:中山大学出版社,1987:132-143.
[3]Shoeb M,Ansari NH,Srivastava SK,et al.4-hydroxynonenal in the pathogenesis and progression of human diseases[J].Current Medicinal Chemistry,2014,21(2):230-237.
[4]Fu SZ,Yin GK,Xin X,et al.Levels of Crotonaldehyde and 4-hydroxy-(E)-2-nonenal and Expression of Genes Encoding Carbonyl-Scavenging Enzyme at Critical Node During Rice Seed Aging[J].Rice Science,2018,25(3):152-160.
[5]A G,AK G,IS S.Changes in oxidative stress enzymes during artificial ageing in cotton(Gossypium hirsutum L.)seeds[J].Journal of Plant Physiology,2003,160(9):1093-1100.
[6]Priestley DA,Leopold C.Tocopherol and Organic Free Radical Levels in Soybean Seeds during Natural and Accelerated Aging[J].Plant Physiology,1980,66(4):715-719.
[7]吴聚兰,周小梅,范玲娟,等.人工老化对大豆种子活力和生理生化特性的影响[J].中国油料作物学报,2011,33(6):582-587.
[8]Tatic'M,Bale2evic'-Tubic'S,or?evie'V,et al.Soybean seed viability and changes of fatty acids content as affected by seed aging[J].African Journal of Biotechnology,2012,(45):10310-10316.
[9]田茜.人工老化对大豆种子线粒体结构功能和抗氧化系统的影响[D].北京:中国农业科学院,2011.
[10]Law MY,Charles SA,Halliwell B.Glutathione and ascorbic acid in spinach(Spinacia oleracea)chloroplasts.The effect of hydrogen peroxide and of Paraquat[J].Biochem J,1983,210(3):899-903.
[11]Ayala A,Mu1oz MF,Argüelles S.Lipid Peroxidation:Production,Metabolism,and Signaling Mechanisms of Malondialdehyde and 4-Hydroxy-2-Nonenal[J].Oxidative Medicine&Cellular Longevity,2014(6):360438.
[12]Selley ML,Bartlett MR,Mcguiness JA,et al.Determination of the lipid peroxidation product trans-4-hydroxy-2-nonenal in biological samples by high-performance liquid chromatography and combined capillary column gas chromatography-negative-ion chemical ionisation mass spectrometry[J].J Chromatogr,1989,488(2):329-340.
[13]Zhang M,Maeda Y,Furihata Y,et al.A mechanism of seed deterioration in relation to the volatile compounds evolved by dry seeds themselves[J].Seed Science Research,1994,4(1):49-56.
[14]Ming Z,Nakamaru Y,Tsuda S,et al.Enzymatic Conversion of Volatile Metabolites in Dry Seeds during Storage[J].Plant&Cell Physiology,1995,36(1):157-164.
[15]董发才,何南峰,张彤,等.人工老化大豆种子膜脂质的过氧化研究[J].河南大学学报(自然科学版),1995(3):91-95.
[16]Asakawa T,Matsushita S.A colorimetric microdetermination of peroxide values utilizing aluminum chloride as the catalyst[J].Lipids,1980,15(11):965-967.
[17]王金龙,李星华.大豆脂氧化酶与种子劣变关系的研究[J].种子,1999,18(6):21-22.
[18]Bale2evic'-Tubic'S,Tatic'M,or?evic'V,et al.Changes in soybean seeds as affected by accelerated and natural aging[J].Romanian Biotechnological Letters,2011,16(6):6740-6747.
[19]董国军,胡兴明,曾大力,等.水稻种子人工老化和自然老化的比较研究[J].浙江农业科学,2004,1(1):27-29.
[20]Vijay D,Dadlani M,Vashisht V.Comparative studies of natural and accelerated ageing in soybean seeds[J].Crop Research,2010,39.
[21]高华伟,满强,武猛,等.耐储藏大豆种质资源的筛选[J].大豆科学,2014,33(1):000006-000012.
[22]Bale2evic'Tubic'S,Tatic'M,or?evic'V,et al.Seed viability of oil crops depending on storage conditions[J].Helia,2010,33(52):153-160.
[23]周小梅,王自霞,乔燕祥.人工老化处理对芝麻种子生理生化特性的影响[J].中国油料作物学报,2008,30(4):460-463.
[24]张瑛,滕斌,吴敬德,等.水稻种子高温高湿人工加速老化试验方法研究[J].中国粮油学报,2010,25(10):8-12.
[25]李稳香,颜启传.杂交水稻自然老化种子与人工老化种子性能差异研究[J].杂交水稻,1997(3):26-28.