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硼是植物必需的微量矿质元素,在植物的生长发育和生理代谢过程中发挥重要作用[1-2] 。但植物对硼元素的适应范围较窄,土壤中有效硼含量过低或过高均会影响植物正常的生长发育[3] ,可能导致叶尖或叶缘失绿、黄化、焦枯[4] ,根系生长受阻、形态改变,植株矮小、死亡等症状。研究发现,硼胁迫抑制了桑树(Morus alba L.)和柑橘(Citrus L.)的株高和根系的生长,降低了叶片的叶绿素含量[5-6] 。并且,硼胁迫会诱导植物组织产生氧化应激损伤,导致脂膜过氧化和活性氧平衡系统改变,在6 mmol·L−1硼酸条件下,苹果砧木EM9(Malus domestica Borkh)的叶和茎中脂氧合酶(lipoxygenase,LOX)活性增强,丙二醛(malondialdehyde,MDA)含量增加,脯氨酸(proline,Pro)含量减少[7] 。氧化应激产物的积累导致氧化还原稳态受损,从而激活植物体内抗氧化系统中保护酶的活性和抗氧化剂含量的变化,硼胁迫导致植物中超氧化物歧化酶(superoxide dismutase,SOD)、过氧化物酶(peroxidase,POD)和过氧化氢酶(catalase,CAT)的活性均增加,并诱导抗坏血酸(ascorbic acid,AsA)含量变化[8] 。当植物处于硼胁迫环境中时其体内的代谢反应也会受到影响,如可溶性蛋白的合成[9-10] 和次生代谢途径相关酶活性的变化[11] 。此外,硼胁迫也会影响植物对其他营养元素的吸收和积累,施加硼营养促进了棉花对氮、磷、钾的吸收,而降低了棉花中钙和镁元素的含量[12] ;而缺硼会降低蚕豆(Vicia faba L.)对磷酸盐的吸收[13] ;研究火炬松(Pinus taeda L.)细胞培养物的生长时,发现硼与钙、镁之间存在着显著的相互作用[14] 。
黑木相思(Acacia melanoxylon R.Br.)属于含羞草亚科(Mimosaceae)金合欢属(Acacia Mill.),是原产于澳大利亚的多年生高大乔木,其适应性广、抗逆性强、速生、材性优良,与根瘤共生固氮,可改良土壤、提升地力、保持水土,具有较好的经济和生态价值[15] 。自20世纪90年代初引入我国,已在广东、广西、海南、福建等华南地区广泛引种栽培。华南地区大部分土壤的有效硼含量较低,达到缺硼(0.25~0.5 mg∙kg−1)或严重缺硼等级(<0.25 mg∙kg−1)[16] ,在林木培育过程中多需施以硼肥,因此,如何避免林木缺硼或硼过量胁迫,是南方人工林培育过程中亟待解决的问题。目前,在杨树(Populus)、桉树(Eucalyptus urophylla S.T. Blake)及针叶树中进行了一些硼营养的研究[17-20],但林木的生长发育对硼胁迫的响应研究仍不够完善。本研究以黑木相思无性系SR17幼苗为材料,分析了不同供硼量条件下幼苗的生长特性、生理生化特征的变化,旨在明确黑木相思对不同供硼量的响应模式,为阐明植物对硼胁迫的抵御机理奠定理论基础,为林木高产、优质栽培提供科学依据。
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不同供硼量条件下,植株的生长表型有所不同(图1)。B0的植株叶片(图1A)比B0.1(图1B)稍黄;而B1和B2呈现出叶缘失绿的表型,且部分叶片黄化脱落(图1C、D)。根系方面,B0形成了白色较粗的主根,侧根也较短而粗(图1E);而B1和B2侧根较细,且随着硼浓度增高而越发褐色(图1G、H)。由此说明,无硼的培养条件使黑木相思幼苗发生缺硼症状,叶片发黄,主根和侧根均加粗,且侧根变短;随着硼浓度的增加,幼苗受到了硼过量的毒害作用,叶缘失绿,老叶黄化脱落,侧根变细,根系褐化。
图 1 不同供硼量对黑木相思幼苗叶和根的影响
Figure 1. Effects of different boric acid content on the leave and roots of A.melanoxylon seedlings
由生长指标测定结果(图2)得出,B0.1植株株高增长量较B0植株株高增长量大;但随着供硼量的继续增加,株高增加量反而减小,B2株高增长量为7.05 cm,仅为B0.1株高增长量的54.9%。根长生长与株高生长情况类似,B0.1根长增长量较B0根长增长量大;而B1和B2的根长增长量仅为B0.1的59.6%和41.7%。随着供硼量的增加,植株叶片数增加量逐渐减少。由此说明,B0.1黑木相思幼苗生长较好;B0形成缺硼表型,叶片发黄,侧根变短,主根和侧根加粗;B1和B2造成硼过量的毒害作用,叶缘失绿,根系褐化,株高、叶片数、根长生长均受到抑制。因此,供硼量是影响黑木相思生长发育的重要因素之一。
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图3表明,B0植株体内叶绿素a和叶绿素b含量均低于B0.1植株;B0.1植株的叶绿素a和叶绿素b含量最高;而B1和B2植株的叶绿素a和叶绿素b的含量逐渐降低。由此推断,0、1和2 mmol·L−1的供硼量均引起黑木相思地上部分叶绿素含量降低,抑制叶片光合作用,从而影响植株的正常生长发育。
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植物体内的可溶性蛋白的含量是反映植物总代谢的重要指标,如图4所示,随着供硼量的增加,黑木相思幼苗体内可溶性蛋白含量越高,植株体内的代谢反应越活跃。AsA是植物细胞中保护叶绿体的重要的抗氧化剂,AsA的含量在B0.1植株体内最高,随着供硼量的增加,AsA的含量急剧降低,说明硼过量条件促进了黑木相思体内的氧化反应造成叶绿体损伤。MDA是过氧脂质分解出的产物之一,其含量可以反应植物体内脂质氧化的水平,在B0、B1和B2 植株体内MDA含量较B0.1植株的高,说明硼胁迫引起黑木相思体内脂质氧化水平升高。Pro是植物抵御逆境时的渗透调节物质之一,用于保持细胞质基质与环境的渗透平衡,防止水分散失,Pro含量在B0、B1和B2 植株体内较高,说明缺硼和硼过量条件造成了黑木相思的渗透胁迫。
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SOD和POD是植物体内氧自由基和H2O2的重要清除剂,保护植物免受氧化反应毒害。如图5所示,在B0、B1和B2幼苗体内SOD和POD的活性较高,推断缺硼和硼过量条件使黑木相思幼苗发生了氧自由基和H2O2的积累。CAT是植物体内清除H2O2的主要酶之一,随着供硼量增加,幼苗体内产生的H2O2越多,CAT的活性越高。PAL是植物体内次生物质苯丙烷类代谢的关键酶,PAL活性在B0、B1和B2幼苗中升高,说明缺硼和硼过量条件促进了黑木相思体内次生代谢物的合成。LOX是一种催化膜脂过氧化的酶,其活性越高使得植物体内的过氧化程度越大,对植株产生毒害作用增加。LOX活性在B0、B1和B2 幼苗体内增加,说明缺硼和硼过量条件使黑木相思幼苗产生了较剧烈的过氧化反应。以上结果表明,缺硼或硼过量条件影响了黑木相思幼苗生化水平的稳定,促进了LOX酶活性的增加,使细胞膜脂产生过氧化反应;促进了SOD、POD、CAT等抗氧化酶和次生代谢物合成酶PAL的活性增加,以抵御过氧化反应的毒害作用,协调控制黑木相思的生长发育。
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在植物体内,不同营养元素之间会产生促进或拮抗的作用,这些相互作用完成植物生理生化过程,调节着植物的营养状况,影响着植物的生长发育。结果表明(图6),植株体内硼元素的含量随着培养液中供硼量的增加而增加,尤其在过量硼供应条件下(B1和B2)发生硼元素的大量积累,分别为B0.1硼含量的10.94倍和12.95倍;随着供硼量的增加,磷和钾元素的含量逐渐增加;而氮和镁元素的含量无明显变化;钙元素在生长状态较好的植株(B0.1 )中的含量较高,随着硼的过量供应,钙含量逐渐降低,B1和B2的钙含量分别较B0.1的降低15.38%和23.08%。由此说明,黑木相思体内硼元素与不同元素之间的相互作用有所不同,硼元素的积累可以促进磷和钾元素的积累,即硼元素与磷和钾元素具有协同作用;硼元素与氮和镁元素无明显相互作用;在硼元素促进植株生长的条件下,可以促进钙元素的积累,但硼过量抑制植株生长时,也抑制了钙元素的积累。
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为了更明确地分析和评价各指标对不同供硼量的响应程度,将具有显著性差异的17个指标进行主成分分析。结果得出(表1),前两个综合指标的贡献率分别为75.14%、19.66%,累计贡献率达94.80%,表明提取前两个主成分可以代表大部分指标的信息。以特征向量系数作为评价标准(表2),第一主成分特征值为12.77,其中钾元素含量、叶片数增长量、株高增长量、LOX活性、AsA含量、钙元素含量、硼元素含量、POD活性、Pro含量、SOD活性、根长增长量、蛋白浓度、PAL活性的荷载量较高;第二主成分特征值为3.34,其中叶绿素b含量的荷载量较高。因此,可取以上14个指标作为评价黑木相思对不同供硼量条件适应性的主要指标。
表 1 综合评价主成分指标系数及贡献率
Table 1. Principal components eigenvectors and cumulative contribution rates
项目
Item主成分1
Prin.1主成分2
Prin.2钾含量 K content −0.98 −0.12 叶片数增长量 Leave number growth −0.98 −0.19 株高增长量 Plant height growth −0.97 0.24 脂氧合酶活性 LOX activity 0.96 0.26 抗坏血酸含量 AsA content −0.95 −0.19 钙含量 Ca content 0.92 0.37 硼含量B content 0.92 −0.31 过氧化物酶活性 POD activity 0.92 −0.23 脯氨酸含量 Pro content 0.91 −0.32 超氧化物歧化酶活性 SOD activity −0.88 −0.36 根长增长量 Root growth −0.88 0.39 蛋白浓度 Protein concentration 0.84 −0.51 苯丙氨酸解氨酶活性 PAL activity 0.81 −0.55 磷含量 P content 0.75 0.37 叶绿素a含量 Chlorophyll a content 0.74 0.67 叶绿素b含量 Chlorophyll b content −0.49 0.87 过氧化氢酶活性 CAT activity 0.67 0.75 特征值 Eigenvalues 12.77 3.34 方差贡献率/%
Variance contribution rate75.14 19.66 累计贡献率/%
Cumulative contribution rate75.14 94.80 表 2 不同供硼量条件下各指标的主成分和隶属函数分析及综合评价
Table 2. Comparison and comprehensive evaluation of the subordinate function values under different boric acid supply
指标
ItemF1 F2 F U(F1) U(F2) D 排名
RankedB0 −1.53 −2.48 −1.73 0.29 0 0.23 3 B0.1 −3.96 1.25 −2.87 0 0.99 0.21 4 B1 1.13 1.29 1.16 0.61 1 0.69 2 B2 4.36 −0.07 3.43 1 0.64 0.92 1 权重Index weight 0.79 0.21 注: F1, F2 分别代表相互独立的综合指标得分,F代表主成分综合得分; U(F1)、U(F2)分别代表各综合指标的隶属函数值,D代表主成分和隶属函数分析的综合评价值
Notes: F1, F2 represent independent comprehensive index, respectively; U(F1)、U(F2) represent the subordinate function values of comprehensive index, respectively; D represents the comprehensive evaluation value of principal component and subordinate function analysis根据主成分特征向量系数及贡献率(表2),获得前两个主成分的因子方程:
F1=−0.27Q1−0.27Q2−0.27Q3+0.27Q4−0.27Q5+0.26Q6+0.26Q7+0.26Q8+0.25Q9−0.25Q10−0.25Q11+0.24Q12+0.23Q13+0.21Q14+0.21Q15−0.14Q16+0.19Q17;
F2=−0.06Q1−0.10Q2+0.13Q3+0.14Q4−0.10Q5+0.20Q6−0.16Q7−0.12Q8−0.17Q9−0.19Q10+0.21Q11−0.27Q12−0.29Q13+0.19Q14+0.35Q15+0.46Q16+0.39Q17(式中Qi为各指标标准化后的值)。
主成分综合得分公式为F=0.79F1 + 0.21F2,分别计算不同供硼量条件下,前两个主成分的综合得分(F),得出黑木相思的生长发育受胁迫程度排序为B2>B1 >B0 >B0.1。另外,利用权重(W)及隶属函数值求得不同供硼量条件下黑木相思幼苗生长发育的综合评价值(D),结果表明(表2),对黑木相思幼苗生长发育的胁迫程度同为B2>B1 >B0>B0.1。因此,黑木相思幼苗相对适宜的供硼量为0.1 mmol·L−1,而 0、1 和2 mmol·L−1的供硼量均影响黑木相思幼苗的生长发育。
硼对黑木相思幼苗生长发育的影响
Effects of Boron on the Growth and Development of Acacia melanoxylon R.Br. seedlings
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摘要:
目的 研究不同供硼量培养条件下,黑木相思生长指标、生理生化特性和营养元素含量的变化,揭示黑木相思对硼胁迫环境的响应模式。 方法 以1月龄黑木相思无性系SR17幼苗为材料,利用含0、0.1、1、2 mmol·L−1硼酸的营养液培养2个月后,测定黑木相思的生长指标、叶绿素含量、生理活性物质含量、氧化还原酶活性和营养元素含量等生理生化指标的变化,基于主成分与隶属函数分析,综合评价各指标对黑木相思响应不同供硼量的贡献率,比较黑木相思幼苗对不同供硼量的适应性。 结果 试验结果表明,0、1和2 mmol·L−1供硼量抑制了黑木相思幼苗株高和根长的增加,降低了地上部分叶绿素的含量,引起叶片失绿、黄化甚至脱落的表型。0 mmol·L−1供硼量使黑木相思的主根和侧根呈白色且增粗,1 、2 mmol·L−1供硼量使根系褐化。0、1、2 mmol·L−1供硼量降低了黑木相思体内抗坏血酸(AsA)的含量,引起了氧化反应;增加了丙二醛(MDA)和脯氨酸(Pro)含量,引起体内脂质氧化水平升高和渗透胁迫;增加了体内超氧化物歧化酶(SOD)、过氧化物酶(POD)、过氧化氢酶(CAT)、脂氧合酶(LOX)和苯丙氨酸解氨酶(PAL)的活性,引发了氧自由基和过氧化氢(H2O2)的积累与清除以及次生代谢物的合成。黑木相思体内硼元素含量增加,磷和钾元素的含量也随之增加,而钙元素含量在 0.1 mmol·L−1供硼量下最高。通过主成分和隶属函数分析,钾元素含量、叶片数增长量、株高增长量、LOX活性、AsA含量、钙元素含量、硼元素含量、POD活性、Pro含量、SOD活性、根长增长量、蛋白浓度、PAL活性及叶绿素b含量14个指标可作为黑木相思生长发育响应硼胁迫的主要指标。 结论 黑木相思幼苗在0.1mmol·L−1供硼量下生长势较好,其生长指标、生理生化特性和营养元素含量在不同供硼量条件下差异显著,黑木相思通过调控植株表型、生理活性物质含量、氧化还原酶活性和营养元素含量响应硼胁迫。 Abstract:Objective To study the changes of growth indicators, physiological and biochemical characteristics, and nutrient element content of Acacia melanoxylon R.Br. under different boron content, and reveal the response mode of A.melanoxylon to boron stress environment. Method The A.melanoxylon clone SR17 1-month-old seedlings were cultured with nutrient solution containing 0 , 0.1, 1, and 2 mmol·L−1 boric acid for 2 months. The changes in the physiological and biochemical indexes, such as growth index, chlorophyll content, physiologically active substances content, oxidoreductase activity, and nutrient element content, were determined under each culture condition. Based on principal component and subordinate function analysis, the contribution rate of each index for A.melanoxylon response to different contribution boron content was comprehensively evaluated, and the stress degree of A.melanoxylon seedlings with different boron supply levels was compared. Result The results showed that 0, 1, and 2 mmol·L−1 boron supply inhibited the increase of plant height and root length, reduced the aboveground chlorophyll content, and caused leaf degreenization, yellowing, and even abruption phenotypes. 0 mM boron made the primary and lateral roots white and thickened, 1 and 2 mmol·L−1 boron made the roots brown. The dosages of 0 , 1, and 2 mmol·L−1 boron reduced the content of ascorbic acid (AsA) and caused an oxidation reaction. The contents of malondialdehyde (MDA) and proline (Pro) were increased, leading to increased lipid oxidation levels and osmotic stress. It increased the activities of superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), lipoxygenase (LOX), and phenylalanine ammonlyase (PAL) in vivo, leading to the accumulation and removal of oxygen-free radicals and hydrogen peroxide (H2O2) as well as the synthesis of secondary metabolites. The increases of boron content of A.melanoxylon promoted the increase of phosphorus and potassium content, and the calcium content was the highest under 0.1 mmol·L−1 boron supply. Through principal component and subordinate function analysis, fourteen indexes, including potassium content, leaf number increase, plant height increase, LOX activity, AsA content, calcium content, boron content, POD activity, Pro content, SOD activity, root length increase, protein concentration, PAL activity, and chlorophyll b content, could be used as the main indexes of A.melanoxylon growth and development response to boron stress. Conclusion A.melanoxylon seedlings grow better under 0.1mmol·L−1 boron supply, and their growth indicators, physiological and biochemical characteristics, and nutrient element content are significantly different under different boron supply conditions. A.melanoxylon responds to boron stress by regulating plant phenotype, the content of physiologically active substances, oxidoreductase activity, and nutrient element content. -
表 1 综合评价主成分指标系数及贡献率
Table 1. Principal components eigenvectors and cumulative contribution rates
项目
Item主成分1
Prin.1主成分2
Prin.2钾含量 K content −0.98 −0.12 叶片数增长量 Leave number growth −0.98 −0.19 株高增长量 Plant height growth −0.97 0.24 脂氧合酶活性 LOX activity 0.96 0.26 抗坏血酸含量 AsA content −0.95 −0.19 钙含量 Ca content 0.92 0.37 硼含量B content 0.92 −0.31 过氧化物酶活性 POD activity 0.92 −0.23 脯氨酸含量 Pro content 0.91 −0.32 超氧化物歧化酶活性 SOD activity −0.88 −0.36 根长增长量 Root growth −0.88 0.39 蛋白浓度 Protein concentration 0.84 −0.51 苯丙氨酸解氨酶活性 PAL activity 0.81 −0.55 磷含量 P content 0.75 0.37 叶绿素a含量 Chlorophyll a content 0.74 0.67 叶绿素b含量 Chlorophyll b content −0.49 0.87 过氧化氢酶活性 CAT activity 0.67 0.75 特征值 Eigenvalues 12.77 3.34 方差贡献率/%
Variance contribution rate75.14 19.66 累计贡献率/%
Cumulative contribution rate75.14 94.80 表 2 不同供硼量条件下各指标的主成分和隶属函数分析及综合评价
Table 2. Comparison and comprehensive evaluation of the subordinate function values under different boric acid supply
指标
ItemF1 F2 F U(F1) U(F2) D 排名
RankedB0 −1.53 −2.48 −1.73 0.29 0 0.23 3 B0.1 −3.96 1.25 −2.87 0 0.99 0.21 4 B1 1.13 1.29 1.16 0.61 1 0.69 2 B2 4.36 −0.07 3.43 1 0.64 0.92 1 权重Index weight 0.79 0.21 注: F1, F2 分别代表相互独立的综合指标得分,F代表主成分综合得分; U(F1)、U(F2)分别代表各综合指标的隶属函数值,D代表主成分和隶属函数分析的综合评价值
Notes: F1, F2 represent independent comprehensive index, respectively; U(F1)、U(F2) represent the subordinate function values of comprehensive index, respectively; D represents the comprehensive evaluation value of principal component and subordinate function analysis -
[1] GARCÍA-SÁNCHEZ F, SIMÓN-GRAO S, MARTÍNEZ-NICOLÁS J J, et al. Multiple stresses occurring with boron toxicity and deficiency in plants[J]. Journal of Hazardous Materials, 2020, 397: 122713. doi: 10.1016/j.jhazmat.2020.122713 [2] 徐芳森, 王运华. 我国作物硼营养与硼肥施用的研究进展[J]. 植物营养与肥料学报, 2017, 23(6):1556-1564. [3] BRDAR-JOKANOVIĆ M. Boron toxicity and deficiency in agricultural plants[J]. International Journal of Mmolecular Sciences, 2020, 21(4): 1424. doi: 10.3390/ijms21041424 [4] MCCAULEY A, JONES C, JACOBSEN J. Plant nutrient functions and deficiency and toxicity symptoms[J]. Nutrient Management Module, 2009, 9: 1-16. [5] SHAH A, WU X, ULLAH A, et al. Deficiency and toxicity of boron: Alterations in growth, oxidative damage and uptake by citrange orange plants[J]. Ecotoxicology and Environmental Safety, 2017, 145: 575-582. doi: 10.1016/j.ecoenv.2017.08.003 [6] TEWARI R K, KUMAR P, SHARMA P N. Morphology and oxidative physiology of boron-deficient mulberry plants[J]. Tree Physiology, 2010, 30(1): 68-77. doi: 10.1093/treephys/tpp093 [7] MMOLASSIOTIS A, SOTIROPOULOS T, TANOU G, et al. Boron-induced oxidative damage and antioxidant and nucleolytic responses in shoot tips culture of the apple rootstock EM 9 (Malus domestica Borkh)[J]. Environmental and Experimental Botany, 2006, 56(1): 54-62. doi: 10.1016/j.envexpbot.2005.01.002 [8] WANG J Z, TAO S T, QI K J, et al. Changes in photosynthetic properties and antioxidative system of pear leaves to boron toxicity[J]. African Journal of Biotechnology, 2011, 10(85): 19693-19700. [9] KELES Y, ÖNCEL I, YENICE N. Relationship between boron content and antioxidant compounds in Citrus leaves taken from fields with different water source[J]. Plant and Soil, 2004, 265(1/2): 345-353. [10] KRUEGER R W, LOVATT C J, ALBERT L S. Metabolic requirement of Cucurbita pepo for boron[J]. Plant physiology, 1987, 83(2): 254-258. doi: 10.1104/pp.83.2.254 [11] RUIZ J M, BRETONES G, BAGHOUR M, et al. Relationship between boron and phenolic metabolism in tobacco leaves[J]. Phytochemistry, 1998, 48(2): 269-272. doi: 10.1016/S0031-9422(97)01132-1 [12] AHMED N, ABID M, AHMAD F, et al. Impact of boron fertilization on dry matter production and mineral constitution of irrigated cotton[J]. Pakistan Journal of Botany, 2011, 43(6): 2903-2910. [13] ROBERTSON G A, LOUGHMAN B C. Reversible effects of boron on the absorption and incorporation of phosphate in Vicia faba L[J]. New Phytologist, 1974, 73(2): 291-298. doi: 10.1111/j.1469-8137.1974.tb04762.x [14] TEASDALE R D, RICHARDS D K. Boron deficiency in cultured pine cells: Quantitative studies of the interaction with Ca and Mg[J]. Plant Physiology, 1990, 93(3): 1071-1077. doi: 10.1104/pp.93.3.1071 [15] MACHADO J S, LOUZADA J L, SANTOS A J A, et al. Variation of wood density and mechanical properties of blackwood (Acacia melanoxylon R. Br. ) [M]. Materials & Design (1980-2015), 2014, 56: 975-980. [16] WANG Y, SHI L, CAO X, et al. Plant boron nutrition and boron fertilization in China[M]. Advances in Plant and Animal Boron Nutrition, 2007: 93-101. [17] Chernobrovkina N P, Robonen E V, Akhmetova G V, et al. Nitrogen and boron dosage effects on arginine accumulation in Scots pine needles[J]. Forests, 2022, 13(3): 417. doi: 10.3390/f13030417 [18] HODECKER B E R, DE BARROS N F, DA SILVA I R, et al. Boron delays dehydration and stimulates root growth in Eucalyptus urophylla (Blake, ST) under osmotic stress[J]. Plant and Soil, 2014, 384(1-2): 185-199. doi: 10.1007/s11104-014-2196-4 [19] KILPELÄINEN J, RÄISÄNEN M, MEHTÄTALO L, et al. The longevity of Norway spruce responses to boron fertilization[J]. Forest Ecology and Management, 2013, 307: 90-100. doi: 10.1016/j.foreco.2013.06.054 [20] YıLDıRıM K, KASıM G Ç. Phytoremediation potential of poplar and willow species in small scale constructed wetland for boron removal[J]. Chemosphere, 2018, 194: 722-736. doi: 10.1016/j.chemosphere.2017.12.036 [21] 邹琦. 植物生理学实验指导[M]. 北京: 中国农业出版社, 2000. [22] KRUGER N J. The Bradford method for protein quantitation[J]. Methods Mol Biol, 1994, 32: 9-15. [23] HEATH R L, PACKER L. Photoperoxidation in isolated chloroplasts: I. Kinetics and stoichiometry of fatty acid peroxidation[J]. Archives of Biochemistry and Biophysics, 1968, 125(1): 189-198. doi: 10.1016/0003-9861(68)90654-1 [24] BATES L S, WALDREN R P, TEARE I D. Rapid determination of free proline for water-stress studies[J]. Plant and Soil, 1973, 39(1): 205-207. doi: 10.1007/BF00018060 [25] TAKAHAMA U, ONIKI T. Regulation of peroxidase-dependent oxidation of phenolics in the apoplast of spinach leaves by ascorbate[J]. Plant and Cell Physiology, 1992, 33(4): 379-387. [26] BEYER JR W F, FRIDOVICH I. Assaying for superoxide dismutase activity: some large consequences of minor changes in conditions[J]. Analytical Biochemistry, 1987, 161(2): 559-566. doi: 10.1016/0003-2697(87)90489-1 [27] RAO M V, PALIYATH G, ORMROD D P. Ultraviolet B and ozone induced biochemical changes in antioxidant enzymes of Arabidopsis thaliana[J]. Plant Physiology, 1996, 110(1): 125-136. doi: 10.1104/pp.110.1.125 [28] MACADAM J W, NELSON C J, SHARP R E. Peroxidase activity in the leaf elongation zone of tall fescue: I. Spatial distribution of ionically bound peroxidase activity in genotypes differing in length of the elongation zone[J]. Plant Physiology, 1992, 99(3): 872-878. doi: 10.1104/pp.99.3.872 [29] WANG Y S, TIAN S P, XU Y. Effects of high oxygen concentration on pro-and anti-oxidant enzymes in peach fruits during postharvest periods[J]. Food Chemistry, 2005, 91(1): 99-104. doi: 10.1016/j.foodchem.2004.05.053 [30] ROSLER J, KREKEL F, AMRHEIN N, et al. Maize phenylalanine ammonia-lyase has tyrosine ammonia-lyase activity[J]. Plant Physiology, 1997, 113(1): 175-179. doi: 10.1104/pp.113.1.175 [31] 何晓群. 多元统计分析(第四) [M]. 北京: 中国人民大学出版社, 2015. [32] WU X, LIU G, RIAZ M, et al. Metabolic changes in roots of trifoliate orange [Poncirus trifoliate (L. ) Raf. ] as induced by different treatments of boron deficiency and resupply[J]. Plant and Soil, 2019, 434(1-2): 217-229. doi: 10.1007/s11104-018-3684-8 [33] GUNES A, SOYLEMEZOGLU G, INAL A, et al. Antioxidant and stomatal responses of grapevine (Vitis vinifera L. ) to boron toxicity[J]. Scientia Horticulturae, 2006, 110(3): 279-284. doi: 10.1016/j.scienta.2006.07.014 [34] SIMÓN I, DÍAZ‐LÓPEZ L, GIMENO V, et al. Effects of boron excess in nutrient solution on growth, mineral nutrition, and physiological parameters of Jatropha curcas seedlings[J]. Journal of Plant Nutrition and Soil Science, 2013, 176(2): 165-174. doi: 10.1002/jpln.201100394 [35] LEHTO T, RUUHOLA T, DELL B. Boron in forest trees and forest ecosystems[J]. Forest Ecology and Management, 2010, 260(12): 2053-2069. doi: 10.1016/j.foreco.2010.09.028 [36] WU X, RIAZ M, YAN L, et al. Boron deficiency in trifoliate orange induces changes in pectin composition and architecture of components in root cell walls[J]. Frontiers in Plant Science, 2017, 8: 1882. doi: 10.3389/fpls.2017.01882 [37] LI M, ZHAO Z, ZHANG Z, et al. Effect of boron deficiency on anatomical structure and chemical composition of petioles and photosynthesis of leaves in cotton (Gossypium hirsutum L.)[J]. Scientific Reports, 2017, 7(1): 4420. doi: 10.1038/s41598-017-04655-z [38] SUPANJANI L K D, LEE K D. Hot pepper response to interactive effects of salinity and boron[J]. Plant Soil and Environment, 2006, 52(5): 227-233. doi: 10.17221/3433-PSE [39] HEGAZI E S, EL-MOTAIUM R A, YEHIA T A, et al. Effect of foliar boron application on boron, chlorophyll, phenol, sugars and hormones concentration of olive (Olea europaea L. ) buds, leaves, and fruits[J]. Journal of Plant Nutrition, 2018, 41(6): 749-765. doi: 10.1080/01904167.2018.1425438 [40] LAMBERS H, PLAXTON W C. Phosphorus: back to the roots[J]. Annual Plant Reviews, 2015, 48: 3-22. [41] WANG M, ZHENG Q, SHEN Q, et al. The critical role of potassium in plant stress response[J]. International Journal of Mmolecular Sciences, 2013, 14(4): 7370-7390. doi: 10.3390/ijms14047370 [42] IRFAN M, ABBAS M, SHAH J A, et al. Interactive effect of phosphorus and boron on plant growth, nutrient accumulation and grain yield of wheat grown on calcareous soil[J]. Eurasian Journal of Soil Science, 2019, 8(1): 17-26. [43] KAYA C, TUNA A L, DIKILITAS M, et al. Supplementary phosphorus can alleviate boron toxicity in tomato[J]. Scientia Horticulturae, 2009, 121(3): 284-288. doi: 10.1016/j.scienta.2009.02.011 [44] ZARE M, ZADEHBAGHERI M, AZARPANAH A. Influence of potassium and boron on some traits in wheat (Triticum aestivum cv. Darab2)[J]. The International Journal of Biotechnology, 2013, 2(8): 141-153. [45] CAKMAK I, RÖMHELD V. Boron deficiency-induced impairments of cellular functions in plants[J]. Plant and Soil, 1997, 193(1/2): 71-83. [46] REEVE E, SHIVE J W. Potassium-boron and calcium-boron relationships in plant nutrition[J]. Soil Science, 1994, 57(1): 1-14. [47] LIU Y, RIAZ M, YAN L, et al. Boron and calcium deficiency disturbing the growth of trifoliate rootstock seedlings (Poncirus trifoliate L. ) by changing root architecture and cell wall[J]. Plant Physiology and Biochemistry, 2019, 144: 345-354. doi: 10.1016/j.plaphy.2019.10.007