ARF基因干扰表达对不同发育阶段和贮藏条件马铃薯酶活性的影响

doi:10.11686/cyxb2015194

草业学报 ›› 2016, Vol. 25 ›› Issue (4): 133-139.DOI: 10.11686/cyxb2015194

ARF基因干扰表达对不同发育阶段和贮藏条件马铃薯酶活性的影响

周香艳^{1, 2}, 张宁¹, 刘柏林^{2, 3}, 裴瑞芳^{1, 3}, 司怀军^{1, 3*, *}, 王蒂^{2, 3}

1.甘肃农业大学生命科学技术学院,甘肃兰州 730070;
2.甘肃农业大学农学院,甘肃兰州 730070;
3.甘肃省作物遗传改良与种质创新重点实验室,甘肃省干旱生境作物学省部共建国家重点实验室培育基地,甘肃兰州 730070

收稿日期:2015-04-14 出版日期:2016-04-20 发布日期:2016-04-20
作者简介:周香艳(1983-),女,内蒙古包头人,在读博士。E-mail:zhxy2008bj@163.com
基金资助:
甘肃农业大学盛彤笙科技创新基金项目(GSAU-STS-1333),国家自然科学基金项目(31160298),甘肃省杰出青年基金项目(1308RJDA011)和甘肃省农业科技创新项目(GNCX-2012-49)资助

Effects of RNAi expression of the ARF gene on enzyme activity at different developmental stages and storage temperatures in potato

ZHOU Xiang-Yan^{1, 2}, ZHANG Ning², LIU Bai-Lin^{2, 3}, PEI Rui-Fang^{1, 3}, SI Huai-Jun^{1, 3, *}, WANG Di^{2, 3}

1.College of Life Science and Technology, Gansu Agricultural University, Lanzhou 730070, China;
2.College of Agronomy, Gansu Agricultural University, Lanzhou 730070, China;
3.Gansu Key Laboratory of Crop Genetic and Germplasm Enhancement, Gansu Provincial Key Laboratory of Aridland Crop Science, Lanzhou 730070, China

Received:2015-04-14 Online:2016-04-20 Published:2016-04-20

摘要/Abstract

摘要： 以前期研究获得的马铃薯栽培品种“甘农薯2号”ADP核糖基化因子(ARF)基因干扰表达转化植株为材料,用qRT-PCR法对转基因植株的ARF基因表达量进行了测定,结果表明,不同发育阶段转基因叶片中ARF基因的相对表达量先降低,生长后期略有增加,表明ARF基因干扰表达量随马铃薯生长发育发生改变。ARF基因干扰表达影响马铃薯叶片中酶活性,不同发育阶段转基因叶片与对照相比,多酚氧化酶(polyphenol oxidase, PPO)活性升高11.61%~27.84%,硝酸还原酶(nitrate reductase, NR)活性提高21.10%~41.32%,磷脂酶D(phospholipase D, PLD)活性降低2.88%~57.64%,蔗糖磷酸合成酶(sucrose phosphate synthase, SPS)活性提高29.00%~39.57%;不同温度(4℃和室温)贮藏的块茎中ARF基因的相对表达量变化趋势一致:均先降低,再升高,但前者较后者ARF基因相对表达量显著降低。室温较4℃贮藏的转基因块茎PPO活性升高30.44%~56.28%,NR活性提高17.41%~40.92%,PLD活性降低24.39%~85.11%,SPS活性提高30.89%~45.78%。室温较4℃贮藏的非转基因块茎PPO活性升高25.11%~70.66%,NR活性提高36.07%~89.62%,PLD活性降低11.35%~72.64%,SPS活性提高27.31%~61.33%。本研究通过探讨ARF基因干扰表达对马铃薯生理生化特性的影响,为进一步研究ARF基因在马铃薯生长发育调控中的作用提供一定的理论基础。

Abstract: ADP ribosylation factor (ARF) gene expression interference in a transformed potato cultivar “Gannongshu 2” was evaluated. Real time fluorescence quantitative polymerase chain reaction (qRT-PCR) showed that the relative expression of the ARF gene varied in the transgenic plants at different developmental stages and increased slightly at latter growth stages, indicating that ARF gene interference expression varied with maturity.Enzyme activity in the leaves was effected by ARF gene interference expression. For leaves of transgenic plants at different development stages, PPO activity increased by 11.61%-27.84%, NR activity increased by 21.10%-41.32%, PLD activity decreased 2.88%-57.64% SPS activity increased by 29.00%-39.57% when compared with the control. Relative expression of the ARF gene in stored tubers under different storage temperatures (room temperature and 4℃)showed the same trend; initially decreasing and then increasing. For transgenic tubers stored at room temperature, the activity of PPO increased by 30.44%-56.28%, NR activity increased by 17.41%-40.92%, PLD activity decreased 24.39%-85.11%, SPS activity increased by 30.89%-45.78% when compared with transgenic tubers stored at 4℃. For non-transgenic tubers stored at room temperature, PPO activity increased by 25.11%-70.66%, NR activity increased by 36.07%-89.62%, PLD activity decreased 11.35%-72.64% and SPS activity increased by 27.31%-61.33%.This study investigated the effect of RNAi expression of ARF gene on physiological and biochemical characteristics in potato to provide a theoretical basis for further study of the regulation role of the ARF gene in potato of different growth and stage of development.

周香艳, 张宁, 刘柏林, 裴瑞芳, 司怀军, 王蒂. ARF基因干扰表达对不同发育阶段和贮藏条件马铃薯酶活性的影响[J]. 草业学报, 2016, 25(4): 133-139.

ZHOU Xiang-Yan, ZHANG Ning, LIU Bai-Lin, PEI Rui-Fang, SI Huai-Jun, WANG Di. Effects of RNAi expression of the ARF gene on enzyme activity at different developmental stages and storage temperatures in potato[J]. Acta Prataculturae Sinica, 2016, 25(4): 133-139.

参考文献

[1] Moss J, Vaughan M. Structure and function of ARF proteins: Activators of cholera toxin and critical components of intracellular vesicular transport processes. Journal of Biological Chemistry, 1995, 270(21): 12327-12330.
[2] Yang Z. Small GTPases: versatile signaling switches in plants. Plant Cell, 2002, 14(suppl 1): 375-388.
[3] Kahn R A, Gilman A G. Purification of a protein cofactor required for ADP-ribosylation of the stimulatory regulatory component of adenylate cyclase by cholera toxin. Journal of Biological Chemistry, 1984, 259(10): 6228-6234.
[4] Hou L, Li J B, Luo X Y, et al . Cloning, expression and characterization of an ADP-ribosylation factor gene from cotton ( Gossypium hirsutum L.). Agronomica Sinica, 2007, 33(8): 1226-1231.
[5] Brown H A, Gutowsk C, Moomaw C R, et al . ADP-ribosylation factor, a small GTP-dependent regulatory protein, stimulates phospholipase D activity. Cell, 1993, 75(6): 1137-1144.
[6] Orci L, Palmer D J, Amherdt M, et al . Coated vesicle assembly in the Golgi requires only coatomer and ARF proteins from the cytosol. Nature, 1993, 364: 732-734.
[7] Gan L, Yu X X, Yu Z, et al . A study on main agronomic traits of yield and quality of clone lines of Solanum tuberosum hybrids F 1 . Acta Prataculturae Sinica, 2013, 22(4): 312-318.
[8] Pei R F, Liu Y, Liu B L, et al . Genetic transformation using RNAi vector containing an ARF gene in potato and effects on physiological characteristics of microtuber. Acta Prataculturae Sinica, 2015, 24(2): 142-147.
[9] Zuk M, Prescha A, Keüpczynaski J, et al . ADP ribosylation factor regulates metabolism and antioxidant capacity of transgenic potato tuber. Journal of Agricultural and Food Chemistry, 2003, 51(1): 288-294.
[10] Kong L F, Zhang J Y, Liu Z P, et al .Cloning of a S-adenosy methionine synthetase gene from Cleistogenes songorica and its expression under drought stress. Acta Prataculturae Sinica, 2013, 22(1): 268-275.
[11] Wang X J, Li Z, Peng Y.The antioxidant enzyme activities and gene expression induced by spermidine in leaves of white clover. Acta Prataculturae Sinica, 2015, 24(4): 140-147.
[12] Zhu G L, Zhong H W, Zhang A Q. Plant Physiology Experiments[M]. Beijing: Beijing University Press, 1990: 37-40.
[13] Zhang Z L, Qu W J. Experimental Guide to the Study of Plant Physiology (Third Edition)[M]. Beijing: Higher Education Press, 2003: 41-43.
[14] Khatoon H, Talat S, Younus H. Phospholipase D from Allium sativum bulbs: a highly active and thermal stable enzyme. International Journal of Biological Macromolecules, 2008, 42(4): 380-385.
[15] Gao J F. Experimental Guide to the Study of Plant Physiology[M]. Beijing: Higher Education Press, 2006: 105-108.
[16] Liu B L, Zhang N, Wen Y K, et al . Identification of differentially expressed genes in potato associated with tuber dormancy release. Molecular Biology Reports, 2012, 39(12): 11277-11287.
[17] Potato Research Institute of Heilongjiang Academy of Agricultural Sciences. Chinese Potato Cultivation[M]. Beijing: Chinese Agricultural Press, 1994: 55-60.
[18] Bi Y R, Liang H G. Studies on polyphenol oxidase in tobacco callu cultures. Biologiae Experimentalis Sinica, 1988, 21(3): 257-263.
[19] Liu L X, Shen F F, Lu H Q, et al . Research advance on sucrose phosphate synthase in sucrose metabolism. Molecular Plant Breeding, 2005, 3(2): 275-281.
[20] Doehlert D C, Huber S C. Regulation of spinach leaf sucrose phosphate synthase by glucose-6-phosphate, inorganic phosphate, and pH. Plant Physiology, 1983, 73(4): 989-994.
[21] Li Y G, Yu Z W, Jiang D, et al . Studies on the dynamic changes of the synthesis of sucrose in the flag leaf and starch in the grain and related enzymes of high-yielding wheat. Agronomica Sinica, 2001, 27(5): 658-664.
[22] Wu X W. Effects of Different Soil Water Content in Early Growth Stage on Quality and Yield of No-Tillage Potato Covered with Straw[D]. Nanning: Guangxi University, 2007: 24.
[23] Pinhero R G, Paliyath G, Yada R Y, et al . Modulation of phospholipase D and lipoxygenase activities during chilling relation to chilling tolerance of maize seedlings. Plant Physiology and Biochemistry, 1998, 36(3): 213-224.
[24] Kim M J, Oh J M, Cheon S H, et al . Thermal inactivation kinetics and application of phospho-and galactolipid-degrading enzymes for evaluation of quality changes in frozen vegetables. Journal of Agricultural and Food Chemistry, 2001, 49(5): 2241-2248.
[25] Yoshida S. Freezing injury and phospholipid degradation in vivo in woody plant cells. III. Effects of freezing on activity of membrane-bound phospholipase D in microsome-enriched membranes. Plant Physiology, 1979, 64(2): 252-256.
[4] 侯磊, 李家宝, 罗小英, 等.棉花ADP-ribosylation factor基因( GhARF 1)的克隆与表达分析. 作物学报, 2007, 33(8): 1226-1231.
[7] 甘霖, 于肖夏, 于卓, 等. 马铃薯杂种F 1 无性株系产量品质等主要农艺性状研究. 草业学报, 2013, 22(4): 312-318.
[8] 裴瑞芳, 刘英, 刘柏林, 等. 马铃薯 ARF 基因RNAi载体的遗传转化及对其试管薯生理特性的影响. 草业学报, 2015, 24(2): 142-147.
[10] 孔令芳, 张吉宇, 刘志鹏, 等. 无芒隐子草SAMSl基因的克隆及干旱胁迫下的表达分析. 草业学报, 2013, 22(1): 268-275.
[11] 王晓娟, 李州, 彭燕. NO参与Spd诱导白三叶抗氧化酶活性及其基因表达. 草业学报, 2015, 24(4): 140-147.
[12] 朱广廉, 钟诲文, 张爱琴. 植物生理学实验[M]. 北京: 北京大学出版社, 1990: 37-40.
[13] 张志良, 瞿伟菁. 植物生理学实验指导(第三版)[M]. 北京: 高等教育出版社, 2003: 41-43.
[15] 高俊风. 植物生理学实验指导[M]. 北京: 高等教育出版社, 2006: 105-108.
[17] 黑龙江省农业科学院马铃薯研究所. 中国马铃薯栽培学[M]. 北京: 中国农业出版社, 1994: 55-60.
[18] 毕玉蓉, 梁厚果. 烟草愈伤组织多酚氧化酶研究. 实验生物学报, 1988, 21(3): 257-263.
[19] 刘凌霄, 沈法富, 卢合全, 等. 蔗糖代谢中蔗糖磷酸合成酶(SPS)的研究进展. 分子植物育种, 2005, 3(2): 275-281.
[21] 李永庚, 于振文, 姜东, 等. 冬小麦旗叶蔗糖和籽粒淀粉合成动态及与其有关的酶活性的研究. 作物学报, 2001, 27(5): 658-664.
[22] 吴晓伟. 生育初期不同土壤水分对免耕稻草覆盖马铃薯产量及品质的影响[D]. 南宁: 广西大学, 2007: 24.

ARF基因干扰表达对不同发育阶段和贮藏条件马铃薯酶活性的影响

Effects of RNAi expression of the ARF gene on enzyme activity at different developmental stages and storage temperatures in potato

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