接种尖镰孢菌对枸杞苯丙烷代谢关键酶及产物的影响

doi:10.11686/cyxb2016024

摘要/Abstract

摘要： 为了阐明抗感枸杞与根腐病病原菌互作过程中苯丙烷代谢途径中关键酶和代谢产物的变化差异与抗病性的关系,以枸杞栽培品种宁杞一号(NingqiⅠ)、宁杞二号(NingqiⅡ),国内野生种中国枸杞、宁夏枸杞,美洲引进野生种L.brevipes、L.exsertum等6种材料为参试材料,采用切根法接种强致病菌尖镰孢菌后统计死亡率,以此筛选出抗病和感病材料,并测定二者0~20 d内苯丙氨酸解氨酶(PAL)、肉桂酸羟化酶(C4H)、4-香豆酰-辅酶A连接酶(4CL)等苯丙烷代谢途径关键酶以及总酚和类黄酮等代谢产物的动态变化。结果表明,6种参试材料接种枸杞根腐病强致病菌尖镰孢菌后,野生材料较栽培材料抗病性强,宁杞一号为感病材料,L.exsertum抗根腐病能力最强;抗病材料L.exsertum苯丙烷代谢关键酶苯丙氨酸解氨酶、肉桂酸羟化酶、4-香豆酰-辅酶A连接酶活性显著高于宁杞一号,初步确定苯丙烷代谢中的苯丙氨酸解氨酶、肉桂酸羟化酶、4-香豆酰-辅酶A连接酶的活性和类黄酮的含量与枸杞材料抗根腐病呈正相关。因此,苯丙氨酸解氨酶、肉桂酸羟化酶、4-香豆酰-辅酶A连接酶活性和类黄酮含量,初步可以作为筛选抗枸杞尖镰孢菌根腐病的生化指标。

Abstract: In this study, the phenylpropanoid pathway of the interaction of wolfberry and root rot pathogens will be discussed. The relationship between wolfberry phenylpropanoid pathway and disease resistance will be clarified. In this study, the experimental materials are cultivated varieties NingqiⅠ, NingqiⅡ; domestic wild species Lyciumchinense,L.barbarum; wild species of America L.brevipes,L.exsertum. The strong pathogenic Fusariumoxysporum isolated from the onset of wolfberry will be inoculation by root cutting method. And the anti-sense wolfbeey will be choose. This 2 kinds of wolfberry as materials to study the change of phenylalanine ammonia lyase, cinnamic acid hydroxylase, 4-hydroxylase p-coumaric acyl-coenzyme A ligase, phenolics and flavonoids in 20 days. The results show that the disease resistance of wild materials are strong than the cultivated material in 6 kinds of test materials which were inoculated with wolfberry root rot strong pathogen F.oxysporum. NingqiⅠis susceptible materials. The resistance to root rot disease of L.exsertum are strongest. The key enzymes of phenylpropanoid pathway like phenylalnine ammonialyase, trans-cinnamic acid-4-hydroxylase, 4-coumarate: coenzyme A ligase activity are significantly higher than Ningqi Ⅰ. Initially identified the phenylalnine ammonialyase, trans-cinnamic acid-4-hydroxylase, 4-coumarate: coenzyme A ligase activity and flavonoid content of phenylpropanoid pathway of test material resistance to root rot disease was positively correlated. Therefore, the phenylalnine ammonialyase, trans-cinnamic acid-4-hydroxylase, 4-coumarate: coenzyme A ligase enzyme activity and flavonoid content can be a screening of anti-Chinese wolfberry Fusarium root rot of biochemical indexes preliminary.

李捷, 冯丽丹, 杨成德, 王有科, 何静, 张宝琳, 陈秀蓉. 接种尖镰孢菌对枸杞苯丙烷代谢关键酶及产物的影响[J]. 草业学报, 2016, 25(5): 87-94.

LI Jie, FENG Li-Dan, YANG Cheng-De, WANG You-Ke, HE Jing, ZHANG Bao-Lin, CHEN Xiu-Rong. EffectsofkeyenzymesandproductsinphenylpropanoidpathwayofLyciuminfectedbyFusariumoxysporum[J]. Acta Prataculturae Sinica, 2016, 25(5): 87-94.

参考文献

[1] Zhang J, Zhang Q, He J M, etal . Estimation and analysis of grass biomass in desert by remote sensing. Acta Ecologica Sinica, 2007, 27(6): 2294-2301.
[2] Luo Q, Li Z, Yan J, etal . Lyciumbarbarum polysaccharides induce apoptosis in human prostate cancer cells and inhibits prostate cancer growth in a xenograft mouse model of human prostate cancer. Journal of Medicinal Food, 2009, 12(4): 695-703.
[3] Zhao R, Li Q W, Xiao B. Effect of Lyciumbarbarum polysaccharide on the improvement of insulin resistance in NIDDM rats. Yakugaku Zasshi, 2005, 125(12): 981-988.
[4] Zhang H F, Wang Y F. The protective effects of Lyciumbararum polysaccharides on alloxan-induced rat pancreatic islets damage. Chinese Journal of Cell Biology, 2005, 27(2): 173-177.
[5] Chen Z, Soo M Y, Srinivasan N, etal . Activation of macrophages by polysaccharide-protein complex from Lyciumbarbarum . Applied Physics Letters, 2015, 106(10): 1116-1122.
[6] Liu Z R. Preliminary study on the withered disease of Chinese wolfberry. Science and Technology of Qinghai Agriculture and Forestry, 1980, 3: 43-45.
[7] Lu Z K, Wang G Z, Zhang L R, etal . Occurrence and control of wolfberry root rot. Acta Phytophylacica Sinica, 1994, 21(3): 249-254.
[8] Li H, Li G Y, Fu J H. Identification of the causing agent of wolfberry root rot in Xinjiang. Journal of Plant Protection, 1998, 25(3): 253-257.
[9] Gao B D, Chen J. Physiological Plant Pathway[M]. Beijing: Science Press, 2006.
[10] Bao G H, Bi Y, Li Y C, etal . Comparasion of phenylpropanoid pathway metabolism in slices of susceptible and resistant potato cultivars inoculated with Fusariumsulphureum . Food Science, 2015, 36(6): 151-156.
[11] Ma J F, Li M Q, Zhang Z H, etal . Study on relationship between phenylalanine ammonialyase (PAL) and resistance to crown and root rot in alfalfa cultivars. Acta Prataculturae Sinica, 2003, 12(4): 35-39.
[12] Song X P, Huang X, Mo F L, etal . Cloning and expression analysis of sugarcane phenylalanin ammonia-lyase ( PAL ) gene. Scientia Agricultura Sinica, 2013, 46(14): 2856-2868.
[13] Gu B G, Zhang Y H, Liu N C, etal . GB/T 17980.88-2004, Fungicides Against Soybean Root Rot[S]. Beijing: Standards Press of China, 2004.
[14] Li H S, Sun Q, Zhao S J, etal . The Principle and Technology of Plant Physiology and Biochemistry Experiment[M]. Beijing: Higher Education Press, 2000: 105-109.
[15] Liu H X, Jiang W B, Bi Y, etal . Postharvest BTH treatment induces resistance of peach ( Prunuspersica L. cv. Jiubao ) fruit to infection by Penicilliumexpansum and enhances activity of fruit defense mechanisms. Postharvest Biology and Technology, 2005, 35(3): 263-269.
[16] Fan C F, Bi Y, Wang Y F, etal . Effect of salicylic acid dipping on postharvest diseases and phenylpropanoid pathway in muskmelon fruits. Scientia Agricultura Sinica, 2012, 45(3): 584-589.
[17] Lamb C J, Rubery P H. A spectrophotometric assay for trans-cinnamic acid 4-hydroxylase activity. Analytical Biochemistry, 1975, 68(2): 554-561.
[18] Zhu M H, Ouyang G C, Xue Y L. Changes of activities of four enzymes (G6PD, PAL, 4CL, PO) and lignin content in immunization induced cucumber. Acta Agriculture Shanghai, 1990, 6(2): 21-26.
[19] Pirie A, Mullins M G. Changes in anthocyanin and phenolics content of grapevine leaf and fruit tissue treated with sucrose, nitrate, and abscisic acid. Plant Physiology, 1976, 58(4): 468-472.
[20] Mauch-Mani B, Slusarenko A J. Production of salicylic acid precursors is a major function of phenylalanine ammonialyase in the resistance of Arabidopsis to Peronosporaparasitica . The Plant Cell, 1996, 8(2): 203-212.
[21] Whetten R, Sederoff R. Lignin biosynthesis. The Plant Cell, 1995, 7(7): 1001-1013.
[22] Xu L F, Zhao H B, Ma F W, etal . Crude toxin isolated from Verturianoshicola on physiological charateristics of in vitro leaves of susceptible and resistant pear. Acta Botanica Boreali-Occidentalia Sinica, 2007, 27(6): 1156-1160.
[23] Li X M, Cheng Z H, Dong Y X. Effects of Alternariaporri (Elliott) Cif. crude toxin on protective enzyme activities and leaf anatomical structure of garlic seedlings. China Vegetables, 2012, 16(1): 68-74.
[24] Xue J, Ma S, Deng X, etal . Prokaryotic expression and reverse catalytic activity characterizations of phenylalanine ammonia-lyase gene ( FtPAL ) from Fagopyrumtataricum . Journal of Agricultural Biotechnology, 2014, 22(1): 64-70.
[25] Liu R F, Liu Q, Zhang F Y, etal . The analysis of differential expression genes for rose early responding to black-spot disease. Acta Horticulturae Sinica, 2015, 42(4): 731-740.
[26] Tian X L, Liu M T, Yang J R. Seed germination and growth of various resistant cucumber seedlings under Fusariumoxysporum crude toxin stress. Chinese Journal of Eco-Agriculture, 2008, 16(6): 1495-1498.
[27] Wang W, Ruan M H, Qiu Y X, etal . Phenylaprapanoid metabolism of sweet potato against Pseudomonassolanacearum . Chinese Journal of Eco-Agriculture, 2009, 17(5): 944-948.
[28] Li L, Zhao Y, Ma J L. Recent progress on key enzymes: PAL, C4H, 4CL of phenylalanine metabolism pathway. China Journal of Bioinformatics, 2007, 5(4): 187-189.
[29] Zhang W Y, Bi Y. Postharvests Diseases and Control of Fruits and Vegetables[M]. Beijing: China Agriculture Press, 1996.
[30] Liang L, Han X M, Zhang Z, etal . Cloning and expression analysis of cinnamate 4-hydroxylase (C4H) reductase gene from Aquilariasinensis . China Journal of Chinese Materia Medica, 2014, 6(10): 705-707.
[31] Chiang V L. Monolignol biosynthesis and genetic engineering of lignin in trees, a review. Environmental Chemistry Letters, 2006, 4(3): 143-146.
[32] Zhao S J, Liu D, Hu Z B. 4-coumarate: Coenzyme a ligase in plant. Plant Physiology Communications, 2006, 42(3): 529-538.
[33] Reimers P J, Leach I E. Race-specific resistance to Xanthomonasoryzae pv. Oryzae conferred by bacterial blight resistance gene Xa-10 in rice ( Oryzaesativa ) involves accumulation of a lignin-like substance in host tissues. Molecular Plant Pathology, 1999, 38(1): 39-55.
[34] Liu M Y, Sun H J, Wang J J, etal . The study on phenol metabolism in sweet potato infected by Ceratocystisfimbriata ellis et halsted. Scientia Agricultura Sinica, 2012, 28(24): 226-230.
[35] Liu P, Shi Y Y, Xue C, etal . Investigation of the resistance to pear valsa canker and the physiological and biochemical characteristics in pyrus germplasm. Acta Botanica Boreali-Occidentalia Sinica, 2014, 34(6): 1164-1172.
[36] Ding Q, Jin S L, Qu Y L, etal . Determination and analysis of antifungal composition content in melon leaves. Journal of Gansu Agricultural University, 2012, 47(4): 57-60, 63.
[1] 张杰,张强,何金梅,等. 荒漠草原牧草生物量的遥感估算及空间分布. 生态学报, 2007, 27(6): 2294-2301.
[6] 刘振荣. 枸杞枯萎病研究初报. 青海农林科技, 1980, 3: 43-45.
[7] 鲁占魁,王国珍,张丽荣,等. 枸杞根腐病的发生及防治研究. 植物保护学报, 1994, 21(3): 249-254.
[8] 李晖,李国英,付建红. 新疆枸杞烂根病病原的鉴定. 植物保护学报, 1998, 25(3): 253-257.
[9] 高必达,陈捷. 生理植物病理学[M]. 北京:科学出版社, 2006.
[10] 包改红,毕阳,李永才,等. 硫色镰刀菌( Fusariumsulphureum )接种对抗病/易感品种马铃薯块茎苯丙烷代谢的影响比较. 食品科学, 2015, 36(6): 151-156.
[11] 马静芳,李敏权,张自和,等. 苯丙氨酸解氨酶与苜蓿种质根和根颈腐烂病抗病性研究. 草业学报, 2003, 12(4): 35-39.
[12] 宋修鹏,黄杏,莫凤连,等. 甘蔗苯丙氨酸解氨酶基因( PAL )的克隆和表达分析. 中国农业科学, 2013, 46(14): 2856-2868.
[13] 顾宝根,张匀华,刘乃炽,等. GB/T 17980. 88-2004,杀菌剂防治大豆根腐病[S]. 北京:中国标准出版社, 2004.
[14] 李合生,孙群,赵世杰,等. 植物生理生化实验原理和技术[M]. 北京:高等教育出版社, 2000: 105-109.
[16] 范存斐,毕阳,王云飞,等. 水杨酸对厚皮甜瓜采后病害及苯丙烷代谢的影响. 中国农业科学, 2012, 45(3): 584-589.
[18] 朱明华,欧阳光察,薛应龙. 黄瓜免疫诱导过程中G6PD、PAL、4CL、PO 活性和木质素含量的变化. 上海农业学报, 1990, 6(2): 21-26.
[22] 徐凌飞,赵竑博,马锋旺,等. 梨黑星菌粗毒素对抗病和感病梨离体叶片生理特性的影响. 西北植物学报, 2007, 27(6): 1156-1160.
[23] 李晓敏,程智慧,董殷鑫. 紫斑病菌粗毒素对大蒜幼苗防御酶活性和叶片解剖结构的影响. 中国蔬菜, 2012, 16(1): 68-74.
[24] 薛军,马双,邓霞,等. 苦荞苯丙氨酸解氨酶基因( FtPAL )的原核表达及其逆向催化酶学性质分析. 农业生物技术学报, 2014, 22(1): 64-70.
[25] 刘瑞峰,刘强,张非亚,等. 月季响应黑斑病的早期差异表达基因分析. 园艺学报, 2015, 42(4): 731-740.
[26] 田雪亮,刘鸣韬,杨家荣. 黄瓜枯萎菌粗毒素对不同抗性黄瓜种子萌发及幼苗胁迫作用研究.中国生态农业学报, 2008, 16(6): 1495-1498.
[27] 王伟,阮妙鸿,邱永祥,等. 甘薯抗薯瘟病的苯丙烷类代谢研究.中国生态农业学报, 2009, 17(5): 944-948.
[28] 李莉,赵越,马君兰. 苯丙氨酸代谢途径关键酶: PAL、C4H、4CL 研究新进展. 生物信息学, 2007, 5(4): 187-189.
[29] 张维一,毕阳. 果蔬采后病害与控制[M]. 北京:中国农业出版社, 1996.
[30] 梁良,韩晓敏,张争,等. 白木香肉桂酸-4-羟基化酶(C4H)基因的克隆及表达分析. 中国中药杂志, 2014, 6(10): 705-707.
[32] 赵淑娟,刘涤,胡之璧. 植物 4-香豆酸:辅酶 A 连接酶. 植物生理学通讯, 2006, 42(3): 529-538.
[34] 刘美艳,孙厚俊,王景景,等. 甘薯块根抗黑斑病酚类物质代谢的研究. 中国农学通报, 2012, 28(24): 226-230.
[35] 刘普,施园园,薛程,等. 梨树腐烂病抗性种质筛选及相关生理生化特性研究. 西北植物学报, 2014, 34(6): 1164-1172.
[36] 丁谦,靳生林,曲亚玲,等. 甜瓜不同品种叶片中抗病成分含量的测定与分析. 甘肃农业大学学报, 2012, 47(4): 57-60, 63.w