南阳烟区施氮量对烤烟品种GS同工酶及氮代谢指标的影响
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摘要
为探明南阳烟区浓香型烤烟品种的适宜施氮量,采用裂区试验设计,通过3个施氮量处理,研究了不同烤烟品种(NC71、云烟99、渠首一号)成熟期谷氨酰胺合成酶(GS)同工酶蛋白表达特性及相关生理指标的变化动态。结果表明:NC71成熟期GS1、GS2和硝酸还原酶(NR)活性较低,谷氨酸脱氢酶(GDH)活性较高,具有较低的氮素同化和再利用能力,同时质外体NH+4浓度和氨气挥发量均较高,因此具有较高的成熟期氮效率,施氮量以纯氮52.5～60.0 kg/hm~2为宜。云烟99成熟期具有较高的GS1活性,衰老降解的氮素被大量再利用,氨气挥发量也较低,因此叶片总氮含量较高,容易贪青晚熟,施氮量以25～30 kg/hm~2最为理想。渠首一号氮效率居于两者之间,适合纯氮40～45 kg/hm~2的中等肥力条件。相关性分析表明,GS同工酶活性与叶片氮代谢生理指标有显著或极显著相关性,可以作为研究烤烟品种氮代谢特征的重要依据。
In order to investigate the appropriate nitrogen application amount for fragrant-flavor tobacco in Nanyang tobacco-growing areas,this paper studied the expression characteristics of glutamine synthase( GS) isozyme protein and the changes of related physiological indexes at mature stages of different cultivars using split zone test design and treated by 3 nitrogen application treatments. The results showed that the activities of GS1,GS2 and NR at maturity stage of NC71 were lower,GDH activity was higher,nitrogen assimilation and reutilization ability were also lower. At the same time,the content of NH+4 in apoplast and ammonia volatilization were both high,thus having high nitrogen efficiency in mature stage,and 52.5 ～ 60.0 kg/hm~2 pure nitrogen application was reasonable. Yunyan 99 had higher GS1 activity at mature stage,nitrogen degraded by aging was reused in a large quantities,and the ammonia volatilization was also low,therefore,the total nitrogen content in leaves was higher,aging could be easily delayed. It was ideal to apply 25 ～ 30 kg/hm~2 of pure nitrogen. The nitrogen efficiency of "Qushou 1"was between the other 2 varieties. It was suit to apply 40 ～ 45 kg/hm~2 of pure nitrogen. Correlation analysis showed that there was a significant or extremely significant correlation between GS isoenzyme activity and leaf nitrogen metabolic index,which could be used as an important basis for studying nitrogen metabolism characteristics of flue-cured tobacco.
In order to investigate the appropriate nitrogen application amount for fragrant-flavor tobacco in Nanyang tobacco-growing areas,this paper studied the expression characteristics of glutamine synthase( GS) isozyme protein and the changes of related physiological indexes at mature stages of different cultivars using split zone test design and treated by 3 nitrogen application treatments. The results showed that the activities of GS1,GS2 and NR at maturity stage of NC71 were lower,GDH activity was higher,nitrogen assimilation and reutilization ability were also lower. At the same time,the content of NH+4 in apoplast and ammonia volatilization were both high,thus having high nitrogen efficiency in mature stage,and 52.5 ～ 60.0 kg/hm~2 pure nitrogen application was reasonable. Yunyan 99 had higher GS1 activity at mature stage,nitrogen degraded by aging was reused in a large quantities,and the ammonia volatilization was also low,therefore,the total nitrogen content in leaves was higher,aging could be easily delayed. It was ideal to apply 25 ～ 30 kg/hm~2 of pure nitrogen. The nitrogen efficiency of "Qushou 1"was between the other 2 varieties. It was suit to apply 40 ～ 45 kg/hm~2 of pure nitrogen. Correlation analysis showed that there was a significant or extremely significant correlation between GS isoenzyme activity and leaf nitrogen metabolic index,which could be used as an important basis for studying nitrogen metabolism characteristics of flue-cured tobacco.
引文
[1]王彦亭.我国烟草育种工作发展思路[J].中国烟草科学,2001,22(4):1-5.Wang Y T. Development thinking of tobacco breeding research in China[J]. Chin. Tob. Sci.,2001,22(4):1-5.
[2]史宏志,韩锦峰.烤烟碳氮代谢几个问题的探讨[J].烟草科技,1998(2):34-36.
[3]刘敬业,冉邦定,李天福,等.烤烟K326成熟期中物质代谢与品质形成关系的研究[J].昆明学院学报,1994(S1):95-103.
[4]秦艳青,李春俭,赵正雄,等.不同供氮方式和施氮量对烤烟生长和氮素吸收的影响[J].植物营养与肥料学报,2007,13(3):436-442.Qin Y Q,Li C J,Zhao Z X,et al.. Effects of rates and methods of N application on growth and N uptake of flue-cured tobacco[J]. Plant Nutr. Fertil. Sci.,2007,13(3):436-442.
[5]赵铭钦,陈秋会,赵明山,等.南阳地区生态条件对不同基因型烤烟品种烟叶化学成分和香气物质含量的影响[J].中国烟草学报,2008,14(1):37-41.Zhao M Q,Chen Q H,Zhao M S,et al.. Effects of ecological condition in Nanyang on chemical components and aroma substances in different genotypic flue-cured tobacco leaves[J].Acta Tab. Sin.,2008,14(1):37-41.
[6]严小龙,张福锁.植物营养遗传[M].北京:中国农业出版社,1997.
[7]段旺军,杨铁钊,刘化冰,等.烟叶氨气补偿点的品种间差异及其与氮素代谢的关系研究[J].植物营养与肥料学报,2011,17(2):419-424.Duan W J,Yang T Z,Liu H B,et al.. Differences in NH3compensation point among tobacco(Nicotiana tabacum L.)cultivars and its relationship with nitrogen metabolism[J].Plant Nutr. Fertil. Sci.,2011,17(2):419-424.
[8] Kichey T,Heumez E,Pocholle D,et al.. Combined agronomic and physiological aspects of nitrogen management in wheat highlight a central role for glutamine synthetase[J]. New Phytol.,2006,169(2):265-278.
[9]邓扬悟.甜瓜谷氨酰胺合成酶基因的克隆和功能研究[D].上海:上海交通大学,博士学位论文,2010.Deng Y W. Cloning and characterization of glutamine synthetase genes in melon[D]. Shanghai:Shanghai Jiao Tong University,Doctor Dissertation,2010.
[10] Purcino AAC,Lima T R,Pinto A C,et al.. Glutamine synthetase response to nitrate in maize genotypes of contrasting nitrogen use efficiency[J]. Maydica,2008,53(53):101.
[11] Wang X,Wei Y,Shi L,et al.. New isoforms and assembly of glutamine synthetase in the leaf of wheat(Triticum aestivum L.)[J]. J. Exp. Bot.,2015,66(21):6827-6834.
[12] Sambrook J,Russell D W,著.黄培堂,等译.分子克隆实验指南精编版[M].北京:化学工业出版社,2007.
[13] O'Neal D,Joy K W. Glutamine synthetase of pea leaves I.Purification, stabilization, and p H optima[J]. Archives Biochem. Biophys.,1973,159:113-122.
[14] Turano F J,Dashner R,Upadhyaya A,et al.. Purification of mitochondrial glutamate dehydrogenase from dark-grown soybean seedlings[J]. Plant Physiol., 1996, 112:1357-1364.
[15]邹琦.植物生理学实验指导[M].北京:中国农业出版社,2000.
[16]武云杰,张小全,段旺军,等.烤烟叶片衰老期氨气挥发特征及其生理调控研究[J].西北植物学报,2012,32(10):2082-2088.Wu Y J,Zhang X Q,Duan W J,et al.. Ammonia volatilization characteristics and its physiological regulation in flue-cured tobacco leaves during senescence[J]. Acta Bot. BorealiOccid. Sin.,2012,32(10):2082-2088.
[17] Zhang Z,Xiong S,Wei Y,et al.. The role of glutamine synthetase isozymes in enhancing nitrogen use efficiency of Nefficient winter wheat[J]. Sci. Rep.,2017,7(1):1-12.
[18]龚明,刘友良,朱培仁.低温下稻苗叶片中蛋白质及游离氨基酸的变化[J].植物生理学报,1989(4):18-22.
[19]韩娜,葛荣朝,赵宝存,等.植物谷氨酰胺合成酶研究进展[J].河北师范大学学报(自然科学版),2004,28(4):407-410.Han N,Ge R C,Zhao B C,et al.. Research development of the glutamine synthetase in plants[J]. J. Hebei Norm. Univ.(Nat. Sci.),2004,28(4):407-410.
[20]刘丽,甘志军,王宪泽.植物氮代谢硝酸还原酶水平调控机制的研究进展[J].西北植物学报,2004,24(7):1355-1361.Liu L,Gan Z J,Wang X Z. Advances of studies on the regulation of nitrate metabolism of plants at nitrate reductase level[J]. Acta Bot. Boreali-Occid. Sin.,2004,24(7):1355-1361.
[21] Nielsen K H,Schjoerring J K. Regulation of apoplastic NH+4concentration in leaves of oil seed rape[J]. Plant Physiol.,1998,118:1361-1368.
[22] Miflin B J,Habash D Z. The role of glutamine synthetase and glutamate dehydrogenase in nitrogen assimilation and possibilities for improvement in the nitrogen utilization of crops[J]. J. Exp. Bot.,2002,53(370):979-987.
[23] Brugiere N,Dubois F,Limami A,et al.. Glutamine synthetase in the phloem plays a major role in controlling proline production[J]. Plant Cell,1999,11:1995-2011.
[24]陈永明,邓海滨,杨键.不同打顶方法对烤烟生长及产量品质影响的研究[A].见:中国烟草学会2005年论文集[C].北京,2005.
[25]杨志晓,刘红峰,张小全,等.不同耐肥性烤烟品种成熟期叶片质外体生理特性研究[J].中国烟草学报,2012,18(6):43-47.Yang Z X,Liu H F,Zhang X Q,et al.. Leaf apoplastic physiological characteristics at maturing stage in flue-cured tobacco cultivars with different tolerance to nitrogen[J]. Acta Tab. Sin.,2012,18(6):43-47.
[26] Gallais A,Hirel B. An approach to the genetics of nitrogen use efficiency in maize[J]. J. Exp. Bot.,2004,55(396):295.
[27] Finnemann H,Gundlach W. Reactor core transient analysis using coupled neutronics and thermal-hydraulic models[A].In:Proceedings of European Nuclear Conference[C].Hamburg Germany,1979.
[28] Masclaux-Daubresse C,Carrayol E,Valadier M H. The two nitrogen mobilisation-and senescence-associated GS1 and GDH genes are controlled by C and N metabolites[J]. Planta,2005,221(4):580-588.
[29]武云杰,杨铁钊,张小全,等.不同烤烟品种烟叶衰老期氨气挥发及其与氮素代谢的相关性[J].中国农业科学,2013,46(19):4027-4034.Wu Y J,Yang T Z,Zhang X Q,et al.. Study on correlation between ammonia volatilization and nitrogen metabolism during the tobacco leaves senescence[J]. Sci. Agric. Sin.,2013,46(19):4027-4034.
[30] Purnell M P,Botella J R. Tobacco isoenzyme 1 of NAD(H)-dependent glutamate dehydrogenase catabolizes glutamate in vivo[J]. Plant Physiol.,2007,143(1):530-539.
[31]曹翠玲,李生秀.供氮水平对小麦生殖生长时期叶片光合速率、NR活性和核酸含量及产量的影响[J].植物学通报,2003,20(3):319-324.Cao C L,Li S X. Effect of nitrogen level on the photosynthetic rate,NR activity and the contents of nucleic acid of wheat leaf in the stage of reproduction[J]. Chin. Bull. Bot.,2003,20(3):319-324.
[2]史宏志,韩锦峰.烤烟碳氮代谢几个问题的探讨[J].烟草科技,1998(2):34-36.
[3]刘敬业,冉邦定,李天福,等.烤烟K326成熟期中物质代谢与品质形成关系的研究[J].昆明学院学报,1994(S1):95-103.
[4]秦艳青,李春俭,赵正雄,等.不同供氮方式和施氮量对烤烟生长和氮素吸收的影响[J].植物营养与肥料学报,2007,13(3):436-442.Qin Y Q,Li C J,Zhao Z X,et al.. Effects of rates and methods of N application on growth and N uptake of flue-cured tobacco[J]. Plant Nutr. Fertil. Sci.,2007,13(3):436-442.
[5]赵铭钦,陈秋会,赵明山,等.南阳地区生态条件对不同基因型烤烟品种烟叶化学成分和香气物质含量的影响[J].中国烟草学报,2008,14(1):37-41.Zhao M Q,Chen Q H,Zhao M S,et al.. Effects of ecological condition in Nanyang on chemical components and aroma substances in different genotypic flue-cured tobacco leaves[J].Acta Tab. Sin.,2008,14(1):37-41.
[6]严小龙,张福锁.植物营养遗传[M].北京:中国农业出版社,1997.
[7]段旺军,杨铁钊,刘化冰,等.烟叶氨气补偿点的品种间差异及其与氮素代谢的关系研究[J].植物营养与肥料学报,2011,17(2):419-424.Duan W J,Yang T Z,Liu H B,et al.. Differences in NH3compensation point among tobacco(Nicotiana tabacum L.)cultivars and its relationship with nitrogen metabolism[J].Plant Nutr. Fertil. Sci.,2011,17(2):419-424.
[8] Kichey T,Heumez E,Pocholle D,et al.. Combined agronomic and physiological aspects of nitrogen management in wheat highlight a central role for glutamine synthetase[J]. New Phytol.,2006,169(2):265-278.
[9]邓扬悟.甜瓜谷氨酰胺合成酶基因的克隆和功能研究[D].上海:上海交通大学,博士学位论文,2010.Deng Y W. Cloning and characterization of glutamine synthetase genes in melon[D]. Shanghai:Shanghai Jiao Tong University,Doctor Dissertation,2010.
[10] Purcino AAC,Lima T R,Pinto A C,et al.. Glutamine synthetase response to nitrate in maize genotypes of contrasting nitrogen use efficiency[J]. Maydica,2008,53(53):101.
[11] Wang X,Wei Y,Shi L,et al.. New isoforms and assembly of glutamine synthetase in the leaf of wheat(Triticum aestivum L.)[J]. J. Exp. Bot.,2015,66(21):6827-6834.
[12] Sambrook J,Russell D W,著.黄培堂,等译.分子克隆实验指南精编版[M].北京:化学工业出版社,2007.
[13] O'Neal D,Joy K W. Glutamine synthetase of pea leaves I.Purification, stabilization, and p H optima[J]. Archives Biochem. Biophys.,1973,159:113-122.
[14] Turano F J,Dashner R,Upadhyaya A,et al.. Purification of mitochondrial glutamate dehydrogenase from dark-grown soybean seedlings[J]. Plant Physiol., 1996, 112:1357-1364.
[15]邹琦.植物生理学实验指导[M].北京:中国农业出版社,2000.
[16]武云杰,张小全,段旺军,等.烤烟叶片衰老期氨气挥发特征及其生理调控研究[J].西北植物学报,2012,32(10):2082-2088.Wu Y J,Zhang X Q,Duan W J,et al.. Ammonia volatilization characteristics and its physiological regulation in flue-cured tobacco leaves during senescence[J]. Acta Bot. BorealiOccid. Sin.,2012,32(10):2082-2088.
[17] Zhang Z,Xiong S,Wei Y,et al.. The role of glutamine synthetase isozymes in enhancing nitrogen use efficiency of Nefficient winter wheat[J]. Sci. Rep.,2017,7(1):1-12.
[18]龚明,刘友良,朱培仁.低温下稻苗叶片中蛋白质及游离氨基酸的变化[J].植物生理学报,1989(4):18-22.
[19]韩娜,葛荣朝,赵宝存,等.植物谷氨酰胺合成酶研究进展[J].河北师范大学学报(自然科学版),2004,28(4):407-410.Han N,Ge R C,Zhao B C,et al.. Research development of the glutamine synthetase in plants[J]. J. Hebei Norm. Univ.(Nat. Sci.),2004,28(4):407-410.
[20]刘丽,甘志军,王宪泽.植物氮代谢硝酸还原酶水平调控机制的研究进展[J].西北植物学报,2004,24(7):1355-1361.Liu L,Gan Z J,Wang X Z. Advances of studies on the regulation of nitrate metabolism of plants at nitrate reductase level[J]. Acta Bot. Boreali-Occid. Sin.,2004,24(7):1355-1361.
[21] Nielsen K H,Schjoerring J K. Regulation of apoplastic NH+4concentration in leaves of oil seed rape[J]. Plant Physiol.,1998,118:1361-1368.
[22] Miflin B J,Habash D Z. The role of glutamine synthetase and glutamate dehydrogenase in nitrogen assimilation and possibilities for improvement in the nitrogen utilization of crops[J]. J. Exp. Bot.,2002,53(370):979-987.
[23] Brugiere N,Dubois F,Limami A,et al.. Glutamine synthetase in the phloem plays a major role in controlling proline production[J]. Plant Cell,1999,11:1995-2011.
[24]陈永明,邓海滨,杨键.不同打顶方法对烤烟生长及产量品质影响的研究[A].见:中国烟草学会2005年论文集[C].北京,2005.
[25]杨志晓,刘红峰,张小全,等.不同耐肥性烤烟品种成熟期叶片质外体生理特性研究[J].中国烟草学报,2012,18(6):43-47.Yang Z X,Liu H F,Zhang X Q,et al.. Leaf apoplastic physiological characteristics at maturing stage in flue-cured tobacco cultivars with different tolerance to nitrogen[J]. Acta Tab. Sin.,2012,18(6):43-47.
[26] Gallais A,Hirel B. An approach to the genetics of nitrogen use efficiency in maize[J]. J. Exp. Bot.,2004,55(396):295.
[27] Finnemann H,Gundlach W. Reactor core transient analysis using coupled neutronics and thermal-hydraulic models[A].In:Proceedings of European Nuclear Conference[C].Hamburg Germany,1979.
[28] Masclaux-Daubresse C,Carrayol E,Valadier M H. The two nitrogen mobilisation-and senescence-associated GS1 and GDH genes are controlled by C and N metabolites[J]. Planta,2005,221(4):580-588.
[29]武云杰,杨铁钊,张小全,等.不同烤烟品种烟叶衰老期氨气挥发及其与氮素代谢的相关性[J].中国农业科学,2013,46(19):4027-4034.Wu Y J,Yang T Z,Zhang X Q,et al.. Study on correlation between ammonia volatilization and nitrogen metabolism during the tobacco leaves senescence[J]. Sci. Agric. Sin.,2013,46(19):4027-4034.
[30] Purnell M P,Botella J R. Tobacco isoenzyme 1 of NAD(H)-dependent glutamate dehydrogenase catabolizes glutamate in vivo[J]. Plant Physiol.,2007,143(1):530-539.
[31]曹翠玲,李生秀.供氮水平对小麦生殖生长时期叶片光合速率、NR活性和核酸含量及产量的影响[J].植物学通报,2003,20(3):319-324.Cao C L,Li S X. Effect of nitrogen level on the photosynthetic rate,NR activity and the contents of nucleic acid of wheat leaf in the stage of reproduction[J]. Chin. Bull. Bot.,2003,20(3):319-324.