人工老化处理对菠菜种子生理生化指标及萌发相关基因表达量的影响
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摘要
为探究菠菜种子老化后活力下降的原因和生理生化指标及基因表达量的变化规律,采用高温高湿人工加速老化的方法处理菠菜种子,并测定菠菜种子人工老化过程中的一系列生理生化指标及萌发相关基因表达量的变化。结果表明:随着老化时间的延长,菠菜种子的根长、发芽势和发芽率,超氧化物歧化酶、过氧化物酶、过氧化氢酶、抗坏血酸过氧化物酶活性以及萌发相关基因(GAI1、GA2ox、RACK1、DGAT、ARP1、ARF、ABA8ox1和VP1)的表达量对比未经老化处理的均逐渐下降;可溶性糖和丙二醛的含量逐渐增加。由于抗氧化酶活性的降低,体内的氧自由基得不到及时的清除,从而导致膜质过氧化作用加剧最后引起生物膜的损伤,这是影响种子萌发和加速种子老化的重要原因之一。
In order to find out the reasons for spinach seed's vigor descent,the changing rule of the physiological and biochemical indexes and the expression of the germination-related genes,this study used the method of high temperature and high humidity artificial accelerated aging to treat spinach seeds.A series of physiological and biochemical indexes and the expression of the germination-related genes were evaluated after artificial aging treatment of spinach seeds.The results showed that the length of root,the germination rate,the germination force,the activity of POD,SOD,CAT,APX and the expression of the germination-related genes(GAI1、GA2 ox、RACK1、DGAT、ARP1、ARF、ABA8 ox1 and VP1)gradually decreased and the content of the soluble sugar and the MDA gradually increased with the prolongation of aging time.Because of the decrease of antioxidant enzyme activity,the free radicals in vivo were not cleared up in time,which leaded to the aggravation of membrane oxidation and the damage of the bio membrane,and it was one of the important reasons caused the reduction of the seed germination and the accelerated aging of the seeds.
In order to find out the reasons for spinach seed's vigor descent,the changing rule of the physiological and biochemical indexes and the expression of the germination-related genes,this study used the method of high temperature and high humidity artificial accelerated aging to treat spinach seeds.A series of physiological and biochemical indexes and the expression of the germination-related genes were evaluated after artificial aging treatment of spinach seeds.The results showed that the length of root,the germination rate,the germination force,the activity of POD,SOD,CAT,APX and the expression of the germination-related genes(GAI1、GA2 ox、RACK1、DGAT、ARP1、ARF、ABA8 ox1 and VP1)gradually decreased and the content of the soluble sugar and the MDA gradually increased with the prolongation of aging time.Because of the decrease of antioxidant enzyme activity,the free radicals in vivo were not cleared up in time,which leaded to the aggravation of membrane oxidation and the damage of the bio membrane,and it was one of the important reasons caused the reduction of the seed germination and the accelerated aging of the seeds.
引文
[1]吴聚兰,周小梅,范玲娟,李君剑,张鑫鑫,张海生.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):582-587Wu J L,Zhou X M,Fan L J,Li J J,Zhang X X,Zhang H S.Effects of artificial aging on seed vigor,physiological and biochemical characteristics of soybean seeds[J].Chinese Journal of Oil Crop Sciences,2011,33(6):582-587(in Chinese)
[2]张兆英.种子劣变机制研究进展[J].沧州师范专科学校学报,2005,21(2):89-90Zhang Z Y.Advances in the research of seed deterioration mechanism[J].Journal of Cangzhou Teacher’s College,2005,21(2):89-90(in Chinese)
[3]Delouche J.Accelerated aging techniques for predicting the relative storability of seed lots[J].Seed Science and Technology,1973,2:427-452
[4]McDonald M B.Seed deterioration:Physiology,repair and assessment[J].Seed Science and Technology,1999,27:177-237
[5]吴晓亮,陈晓玲,辛萍萍,张志娥,陶澜,卢新雄.超干燥处理对豌豆种子抗氧化系统酶及热稳定蛋白的影响[J].园艺学报,2006,33(3):523-528Wu X L,Chen X L,Zhang Z E,Tao L,Lu X X.Effect of ultradrying on antioxidant enzymes and heat-stable protein of Pea seeds[J].Acta Horticulturae Sinica,2006,33(3):523-528(in Chinese)
[6]崔鸿文,王飞.黄瓜种子人工老化过程中某些生理生化规律研究[J].西北农林科技大学学报:自然科学版,1992,20(1):51-54Cui H W,Wang F.A study on the physiological and biochemical regularities during artificial aging of cucumber seeds[J].Journal of Northwest Sci-Tech University of Agriculture and Forestry,1992,20(1):51-54(in Chinese)
[7]薄鹏飞,孙秀玲,孙同虎,杜希华,俞慈英.NaCl胁迫对海滨木槿抗氧化系统和渗透调节的影响[J].西北植物学报,2008,28(1):113-118Bo P F,Sun X L,Sun T H,Du X H,Yu C Y.Antioxidative system and osmotic regulation in seedlings of annual hibiscus hamabo Sieb et Zucc under salt stress[J].Acta Botanica Boreali-Occidentalia Sinica,2008,28(1):113-118(in Chinese)
[8]王煜,田廷亮,扶惠作,殷荣华,钱秀珍.油菜种子老化过程中的生理生化变异[J].中国油料,1994,16(3):11-14Wang H,Tian Y L,Fu H Z,Yin R H,Qian X Z.Physiological and biochemical variation in the aging process of Rape seed[J].Chinese Journal of Oil Crop Sciences,1994,16(3):11-14(in Chinese)
[9]孙永伟,聂丽娜,马有志,徐兆师,夏兰琴.小麦穗发芽抗性相关Vp1基因启动子的分离及功能验证[J].作物学报,2011,37(10):1743-1751Sun Y W,Nie L N,Ma Y Z,Xu Z S,Xia L Q.Cloning and functional analysis of viviparous-1promoter in wheat[J].Acta Agronomica Sinica,2011,37(10):1743-1751(in Chinese)
[10]盛慧,秦智伟,李文滨,周秀艳,武涛,辛明.黄瓜生长素反应因子(ARF)家族鉴定及表达特异性分析[J].中国农业科学,2014,47(10):1985-1994Sheng H,Qin Z W,Li W B,Zhou X Y,Wu T,Xin M.Genomewide identification and expression analysis of auxin response factor(ARF)family in Cucumber[J].Scientia Agricultura Sinica,2014,47(10):1985-1994(in Chinese)
[11]李成.生长素抑制蛋白ARP1对烟草生长和抗病性的交叉调控作用[D].南京:南京农业大学,2015Li C.Auxin-repressed protein ARP1regulates plant growth and disease resistance crosstalk in Tobacco[D].Nanjing:Nanjing Agricultural University,2015(in Chinese)
[12]张海军.褪黑素处理对氯化钠胁迫下黄瓜种子萌发的影响及其分子机制[D].北京:中国农业大学,2016Zhang H J.Regulation of melatonin on seed germination under salt stress in cucumber[D].Beijing:China Agricultural University,2016(in Chinese)
[13]陈秀玲,刘思源,孙婷,刘智琛,陶金宝,赵迎新,王傲雪.番茄YABBY基因家族的生物信息学分析[J].东北农业大学学报,2017,48(10):11-19Chen X L,Liu S Y,Sun T,Liu Z C,Tao J B,Zhao Y X,Wang A X.Bioinformatics analysis on YABBYgene family in tomato[J].Journal of Northeast Agricultural University,2017,48(10):11-19(in Chinese)
[14]姚远.OsABA8ox2基因对水稻叶片水分保持的影响及机理研究[D].哈尔滨:哈尔滨师范大学,2015Yao Y.Study on the effect of water retention and mechanism of rice by OsABA8ox2 overexpression[D].Harbin:Harbin Normal University,2015(in Chinese)
[15]李大红,张冬平,曹丹丹,梁建生.植物RACK1蛋白研究进展[J].植物学报,2011,46(2):224-232Li D H,Zhang D P,Cao D D,Liang J S.Research advances in plant RACK1proteins[J].Chinese Bulletin of Botany,2011,46(2):224-232(in Chinese)
[16]崔金秋.芍药种子下胚轴休眠与萌发相关调控基因PlGAI1的克隆及功能分析[D].沈阳:沈阳农业大学,2016Cui J Q.Cloning and functional analysis of the PLGAI1 gene related to the dormancy and germination of the seeds in Paeonia lactiflora pall[D].Shenyang:Shenyang Agricultural University,2016(in Chinese)
[17]唐桂英,柳展基,单雷.二酰基甘油酰基转移酶(DGAT)研究进展[J].中国油料作物学报,2010,32(2):320-328Tang G Y,Liu Z J,Shan L.Progress of diacylglycerol acyltransferase(DGAT)study[J].Chinese Journal of Oil Crop Sciences,2010,32(2):320-328(in Chinese)
[18]Ellis R H,Hong,T D,Roberts E H.A Low-postire-content limit to logarithmic relations between seed moisture content and longevity[J].Annals of Botany,1988,61:405-408
[19]Ellis R H,Hong,T D,Roberts E H.Moisture content and the longevity of seeds of phaseolus vulgaris[J].Annals of Botany,1990,66:341-348
[20]Rajjou L,Lovigny Y,Groot S P C,Belghazi M,Job C,Job D.Proteome-wide characterization of seed aging in Arabidopsis:Acomparison between artificial and natural aging protocols[J].Plant Physiology,2008,148:620-641
[21]Roberts E H.Loss of viability:Chromosomal and genetical aspects[J].Seed Science and Technology,1973(1):515-527
[22]孙春青,杨伟,戴忠良,潘跃平.人工老化处理对结球甘蓝种子生理生化特性的影响[J].西北植物学报,2012,32(8):1615-1620Sun C Q,Yang W,Dai Z L,Pan Y P.Physiological and biochemical characteristics of‘Dongsheng’Cabbage seeds during artificial aging[J].Acta Botanica Boreali-Occidentalia Sinica,2012,32(8):1615-1620(in Chinese)
[23]刘明久,王铁固,陈士林,王春虎,赵新亮.玉米种子人工老化过程中生理特性与种子活力的变化[J].核农学报,2008,22(4):510-513Liu M J,Wang T G,Chen S L,Wang C H,Zhao X L.Physioloycial and seed vigour changes of Maize seeds during artificial aging course[J].Journal of Nuclear Agricultural Sciences,2008,22(4):510-513(in Chinese)
[24]薄丽萍.不结球白菜种子老化衰变生理基础的研究[D].南京:南京农业大学,2011Bo L P.Study on physiological basis of seed aging in nonheading Chinese Cabbage[D].Nanjing:Nanjing Agricultural University,2011(in Chinese)
[25]Hodges D,DeLong J,Forney C,Prange R.Improving the thiobarbituric acid-reactive-substances assay for estimating lipid peroxidation in plant tissues containing anthocyanin and other interfering compounds[J].Planta,1999,207:604-611
[26]李泽琴,李静晓,张根发.植物抗坏血酸过氧化物酶的表达调控以及对非生物胁迫的耐受作用[J].遗传,2013,35(1):45-54Li Z Q,Li X J,Zhang G F.Expression regulation of plant ascorbate peroxidase and its tolerance to abiotic stresses[J].Hereditas,2013,35(1):45-54(in Chinese)
[27]李颜,王倩.大葱种子人工老化与膜脂过氧化的研究[J].种子,2007,26(3):27-30Li Y,Wang Q.The artificial aging of welsh onion seed and its relation to membrane lipid peroxidation[J].Seed,2007,26(3):27-30(in Chinese)
[28]周建萍,乔燕祥,穆志新,王果萍.大豆种子老化过程中活力指标的研究[J].山西农业科学,2007,35(3):33-35Zhou J P,Qiao Y X,Mu Z X,Wang G P.Study on vitality index of soybean seed in aging process[J].Journal of Shanxi Agricultural Sciences,2007,35(3):33-35(in Chinese)
[29]顾炳朝,岳绪国,杨军,唐泽庆.油菜种子老化处理后的生理生化变化[J].福建农业学报,2013,28(4):339-342Gu B C,Yue X G,Yang J,Tang Z Q.Physiological and biochemical characteristics of artificially aged rapeseeds[J].Fujian Journal of Agricultural Sciences,2013,28(4):339-342(in Chinese)
[2]张兆英.种子劣变机制研究进展[J].沧州师范专科学校学报,2005,21(2):89-90Zhang Z Y.Advances in the research of seed deterioration mechanism[J].Journal of Cangzhou Teacher’s College,2005,21(2):89-90(in Chinese)
[3]Delouche J.Accelerated aging techniques for predicting the relative storability of seed lots[J].Seed Science and Technology,1973,2:427-452
[4]McDonald M B.Seed deterioration:Physiology,repair and assessment[J].Seed Science and Technology,1999,27:177-237
[5]吴晓亮,陈晓玲,辛萍萍,张志娥,陶澜,卢新雄.超干燥处理对豌豆种子抗氧化系统酶及热稳定蛋白的影响[J].园艺学报,2006,33(3):523-528Wu X L,Chen X L,Zhang Z E,Tao L,Lu X X.Effect of ultradrying on antioxidant enzymes and heat-stable protein of Pea seeds[J].Acta Horticulturae Sinica,2006,33(3):523-528(in Chinese)
[6]崔鸿文,王飞.黄瓜种子人工老化过程中某些生理生化规律研究[J].西北农林科技大学学报:自然科学版,1992,20(1):51-54Cui H W,Wang F.A study on the physiological and biochemical regularities during artificial aging of cucumber seeds[J].Journal of Northwest Sci-Tech University of Agriculture and Forestry,1992,20(1):51-54(in Chinese)
[7]薄鹏飞,孙秀玲,孙同虎,杜希华,俞慈英.NaCl胁迫对海滨木槿抗氧化系统和渗透调节的影响[J].西北植物学报,2008,28(1):113-118Bo P F,Sun X L,Sun T H,Du X H,Yu C Y.Antioxidative system and osmotic regulation in seedlings of annual hibiscus hamabo Sieb et Zucc under salt stress[J].Acta Botanica Boreali-Occidentalia Sinica,2008,28(1):113-118(in Chinese)
[8]王煜,田廷亮,扶惠作,殷荣华,钱秀珍.油菜种子老化过程中的生理生化变异[J].中国油料,1994,16(3):11-14Wang H,Tian Y L,Fu H Z,Yin R H,Qian X Z.Physiological and biochemical variation in the aging process of Rape seed[J].Chinese Journal of Oil Crop Sciences,1994,16(3):11-14(in Chinese)
[9]孙永伟,聂丽娜,马有志,徐兆师,夏兰琴.小麦穗发芽抗性相关Vp1基因启动子的分离及功能验证[J].作物学报,2011,37(10):1743-1751Sun Y W,Nie L N,Ma Y Z,Xu Z S,Xia L Q.Cloning and functional analysis of viviparous-1promoter in wheat[J].Acta Agronomica Sinica,2011,37(10):1743-1751(in Chinese)
[10]盛慧,秦智伟,李文滨,周秀艳,武涛,辛明.黄瓜生长素反应因子(ARF)家族鉴定及表达特异性分析[J].中国农业科学,2014,47(10):1985-1994Sheng H,Qin Z W,Li W B,Zhou X Y,Wu T,Xin M.Genomewide identification and expression analysis of auxin response factor(ARF)family in Cucumber[J].Scientia Agricultura Sinica,2014,47(10):1985-1994(in Chinese)
[11]李成.生长素抑制蛋白ARP1对烟草生长和抗病性的交叉调控作用[D].南京:南京农业大学,2015Li C.Auxin-repressed protein ARP1regulates plant growth and disease resistance crosstalk in Tobacco[D].Nanjing:Nanjing Agricultural University,2015(in Chinese)
[12]张海军.褪黑素处理对氯化钠胁迫下黄瓜种子萌发的影响及其分子机制[D].北京:中国农业大学,2016Zhang H J.Regulation of melatonin on seed germination under salt stress in cucumber[D].Beijing:China Agricultural University,2016(in Chinese)
[13]陈秀玲,刘思源,孙婷,刘智琛,陶金宝,赵迎新,王傲雪.番茄YABBY基因家族的生物信息学分析[J].东北农业大学学报,2017,48(10):11-19Chen X L,Liu S Y,Sun T,Liu Z C,Tao J B,Zhao Y X,Wang A X.Bioinformatics analysis on YABBYgene family in tomato[J].Journal of Northeast Agricultural University,2017,48(10):11-19(in Chinese)
[14]姚远.OsABA8ox2基因对水稻叶片水分保持的影响及机理研究[D].哈尔滨:哈尔滨师范大学,2015Yao Y.Study on the effect of water retention and mechanism of rice by OsABA8ox2 overexpression[D].Harbin:Harbin Normal University,2015(in Chinese)
[15]李大红,张冬平,曹丹丹,梁建生.植物RACK1蛋白研究进展[J].植物学报,2011,46(2):224-232Li D H,Zhang D P,Cao D D,Liang J S.Research advances in plant RACK1proteins[J].Chinese Bulletin of Botany,2011,46(2):224-232(in Chinese)
[16]崔金秋.芍药种子下胚轴休眠与萌发相关调控基因PlGAI1的克隆及功能分析[D].沈阳:沈阳农业大学,2016Cui J Q.Cloning and functional analysis of the PLGAI1 gene related to the dormancy and germination of the seeds in Paeonia lactiflora pall[D].Shenyang:Shenyang Agricultural University,2016(in Chinese)
[17]唐桂英,柳展基,单雷.二酰基甘油酰基转移酶(DGAT)研究进展[J].中国油料作物学报,2010,32(2):320-328Tang G Y,Liu Z J,Shan L.Progress of diacylglycerol acyltransferase(DGAT)study[J].Chinese Journal of Oil Crop Sciences,2010,32(2):320-328(in Chinese)
[18]Ellis R H,Hong,T D,Roberts E H.A Low-postire-content limit to logarithmic relations between seed moisture content and longevity[J].Annals of Botany,1988,61:405-408
[19]Ellis R H,Hong,T D,Roberts E H.Moisture content and the longevity of seeds of phaseolus vulgaris[J].Annals of Botany,1990,66:341-348
[20]Rajjou L,Lovigny Y,Groot S P C,Belghazi M,Job C,Job D.Proteome-wide characterization of seed aging in Arabidopsis:Acomparison between artificial and natural aging protocols[J].Plant Physiology,2008,148:620-641
[21]Roberts E H.Loss of viability:Chromosomal and genetical aspects[J].Seed Science and Technology,1973(1):515-527
[22]孙春青,杨伟,戴忠良,潘跃平.人工老化处理对结球甘蓝种子生理生化特性的影响[J].西北植物学报,2012,32(8):1615-1620Sun C Q,Yang W,Dai Z L,Pan Y P.Physiological and biochemical characteristics of‘Dongsheng’Cabbage seeds during artificial aging[J].Acta Botanica Boreali-Occidentalia Sinica,2012,32(8):1615-1620(in Chinese)
[23]刘明久,王铁固,陈士林,王春虎,赵新亮.玉米种子人工老化过程中生理特性与种子活力的变化[J].核农学报,2008,22(4):510-513Liu M J,Wang T G,Chen S L,Wang C H,Zhao X L.Physioloycial and seed vigour changes of Maize seeds during artificial aging course[J].Journal of Nuclear Agricultural Sciences,2008,22(4):510-513(in Chinese)
[24]薄丽萍.不结球白菜种子老化衰变生理基础的研究[D].南京:南京农业大学,2011Bo L P.Study on physiological basis of seed aging in nonheading Chinese Cabbage[D].Nanjing:Nanjing Agricultural University,2011(in Chinese)
[25]Hodges D,DeLong J,Forney C,Prange R.Improving the thiobarbituric acid-reactive-substances assay for estimating lipid peroxidation in plant tissues containing anthocyanin and other interfering compounds[J].Planta,1999,207:604-611
[26]李泽琴,李静晓,张根发.植物抗坏血酸过氧化物酶的表达调控以及对非生物胁迫的耐受作用[J].遗传,2013,35(1):45-54Li Z Q,Li X J,Zhang G F.Expression regulation of plant ascorbate peroxidase and its tolerance to abiotic stresses[J].Hereditas,2013,35(1):45-54(in Chinese)
[27]李颜,王倩.大葱种子人工老化与膜脂过氧化的研究[J].种子,2007,26(3):27-30Li Y,Wang Q.The artificial aging of welsh onion seed and its relation to membrane lipid peroxidation[J].Seed,2007,26(3):27-30(in Chinese)
[28]周建萍,乔燕祥,穆志新,王果萍.大豆种子老化过程中活力指标的研究[J].山西农业科学,2007,35(3):33-35Zhou J P,Qiao Y X,Mu Z X,Wang G P.Study on vitality index of soybean seed in aging process[J].Journal of Shanxi Agricultural Sciences,2007,35(3):33-35(in Chinese)
[29]顾炳朝,岳绪国,杨军,唐泽庆.油菜种子老化处理后的生理生化变化[J].福建农业学报,2013,28(4):339-342Gu B C,Yue X G,Yang J,Tang Z Q.Physiological and biochemical characteristics of artificially aged rapeseeds[J].Fujian Journal of Agricultural Sciences,2013,28(4):339-342(in Chinese)
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