【作者】 王艳；

【导师】 何睿；

【作者基本信息】 河南农业大学 ， 植物保护， 2025， 硕士

【摘要】 农田镉污染导致了小麦产量降低和品质下降,已对全球粮食安全与人类健康构成严重威胁。纳米材料因其独特的物理化学性质被广泛应用于改善农作物生长、增强农作物抗重金属胁迫性能。本课题组前期羟基化纳米碳（富勒醇,Fol）提高玉米铜耐受性的研究方面。在此基础上,本论文继续开展Fol抵御小麦幼苗镉胁迫的作用机制研究。通过离体实验测试Fol的抗氧化性能、Fol与Cd²⁺的相互作用,研究Fol对镉胁迫下小麦生长表型、生理生化、分子生物学层面的影响,揭示小麦幼苗响应镉胁迫的“体外”和“体内”协同调控的“金属阻控-抗氧化防御-形态修复”机理。具体研究结果如下:（1）通过NBT、TMB比色实验确证了离体下Fol具有直接清除H₂O₂和O₂^–·能力,且Fol相较于POD与SOD具有高稳定性。由此发现了Fol在一定程度可作为小麦体内POD与SOD的补充剂,可潜在地直接清除由镉胁迫造成的过量累积的H₂O₂和O₂^–·。（2）Cd²⁺能够诱导Fol形成水不溶/水溶性的Cd²⁺@Fol纳米复合物,通过表征分析Fol与Cd²⁺的相互作用。Fol作用于镉胁迫的小麦幼苗时,一方面,Fol可在植株体内复合Cd²⁺,形成水溶性Cd²⁺@Fol-1,具有移动性和易吸收的特点,降低了体内游离Cd²⁺的含量。另一方面,Fol体外固定Cd²⁺形成水不溶的Cd²⁺@Fol-2易进一步絮凝沉降导致部分Cd²⁺被固定,难以被吸收进入小麦幼苗。两种作用协同将显著减少镉胁迫下小麦整株镉积累及体内游离Cd²⁺含量,从而有效降低幼苗的镉毒性。（3）通过对小麦进行水培实验发现了Fol能有效改善镉胁迫下幼苗的生长。相对于单独镉胁迫处理,Fol-Cd共暴露处理使小麦叶绿素和类胡萝卜素含量均呈显著增加趋势。Fol还增加小麦Cd胁迫下的GSH含量,降低AsA、H₂O₂和MDA含量,增强GR、APX、SOD、CAT酶活性,通过维持胞内氧化还原平衡,提高了幼苗抗镉胁迫性能。ICP-MS测试证实,Fol处理降低了Cd吸收效率,减少小麦幼苗各组织内积累的Cd含量。此外,Fol还能有效诱导根际有机酸-镉复合物的形成,这可部分地钝化镉,增强幼苗耐受而抵御镉毒害。（4）在分子生物学层面上,对小麦金属转运蛋白的基因表达量进行了测定,发现了Fol处理降低了游离Cd²⁺对幼苗根部TaNramp5、TaZIP8高表达量的诱导,从而限制幼苗的Cd²⁺吸收。Fol处理还降低了TaHIPP3、TaHIPP20表达量,使得根部捕获在小麦根部胞质溶胶的Cd²⁺减少。同时,Fol处理上调TaZIP9表达量,可促进Cd²⁺在小麦幼苗体内的转运。Fol处理使根部TaHMA2和TaHMA4表达量上调,可促进镉从根部到芽的运输。总之,Fol可抑制镉胁迫下小麦幼苗根部Cd²⁺积累,降低了Cd²⁺的吸收效率,限制小麦对Cd²⁺吸收,促进Cd²⁺的转运,从而助于镉胁迫下幼苗的解毒。同时,通过促进光合作用,调控抗氧化系统,加强根际毒性较低的有机酸-镉复合物的形成,提高幼苗抗逆性而抵御镉胁迫。更多还原

【Abstract】 Cadmium（Cd）contamination in farmland has led to reduced wheat yield and quality,posing a serious threat to global food security and human health.Due to their unique physicochemical properties,nanomaterials have been widely used to improve crop growth and enhance resistance to heavy metal stress.Our research group has made significant progress in using hydroxylated nanocarbon（fullerenol,Fol）to enhance copper tolerance in maize.Building on this foundation,this study further investigates the mechanism by which Fol alleviates cadmium stress in wheat seedlings.The antioxidant performance of Fol and the interaction between Fol and Cd²⁺were tested through in vitro experiments.The effects of Fol on the growth phenotype,physiological and biochemical,and molecular biological aspects of wheat under cadmium stress were studied,revealing the"metal inhibition-antioxidant defense-morphological repair"mechanism of"in vitro"and"in vivo"coordinated regulation of wheat seedlings response to cadmium stress.The specific findings are as follows:（1）Through NBT and TMB colorimetric assays,it was confirmed that Fol can directly scavenge H₂O₂and O₂^–·in vitro,exhibiting higher stability compared to POD and SOD enzymes.Fol exhibits higher stability compared to POD and SOD.Fol can potentially serve as a supplement to POD and SOD in wheat,directly scavenging the excessive accumulation of H₂O₂and O₂^–·under Cd stress.（2）Cd²⁺can induce the formation of water-insoluble/soluble Cd²⁺@Fol nanocomplexes.Characterization analysis revealed the interaction between Fol and Cd²⁺.When applied to Cd-stressed wheat seedlings,Fol exhibits dual mechanisms:In vivo,Fol complexes with Cd²⁺to form water-soluble Cd²⁺@Fol-1,which is mobile and easily absorbed,reducing free Cd²⁺levels.In vitro,Fol immobilizes Cd²⁺,forming water-insoluble Cd²⁺@Fol-2 that tends to flocculate and precipitate,partially fixing Cd²⁺and preventing its uptake by seedlings.These synergistic effects significantly reduce total Cd accumulation and free Cd²⁺content in wheat,thereby alleviating Cd toxicity.（3）Hydroponic experiments of wheat seedlings show that Fol significantly improves seedling growth under Cd stress.Compared to Cd stress,coexposure of Fol and cadmium chloride（Fol-Cd）leads to a significant increase in chlorophyll and carotenoid contents in wheat leaves.It also increases GSH content,reduces H₂O₂,AsAand MDA levels,enhances the enzyme activities of GR,APX,SOD,and CAT,and regulates the antioxidant system to maintain intracellular redox balance,thereby improving seedling resistance to Cd stress;Reduce Cd absorption efficiency and decrease the accumulated Cd contents in shoots and roots of wheat seedlings.Additionally,Fol effectively induces the formation of rhizosphere organic acid-Cd complexes,which partially passivate Cd and enhance seedling tolerance to Cd toxicity.（4）At the molecular biology level,by measuring the expression levels of genes related to wheat metal transporters,it was found that Fol treatment reduced the induction of high expression of TaNramp5 and TaZIP8 in the roots of seedlings by free Cd²⁺,thereby limiting the absorption of Cd²⁺by seedlings.Additionally,Fol treatment decreased the expression levels of TaHIPP3 and TaHIPP20,resulting in a reduction of Cd²⁺captured in the cytosol of wheat roots.Meanwhile,Fol treatment upregulated the expression of TaZIP9,which promoted the transport of Cd²⁺within wheat seedlings.Furthermore,Fol treatment increased the expression levels of TaHMA2 and TaHMA4 in roots,facilitating the transport of cadmium from roots to shoots.In conclusion,Fol can inhibit the accumulation of Cd²⁺in the roots of wheat seedlings under cadmium stress,reduce the absorption efficiency of Cd²⁺,limit the absorption of Cd²⁺by wheat,and promote the transport of Cd²⁺,thereby facilitating the detoxification of seedlings under cadmium stress.Meanwhile,by promoting photosynthesis,regulating the antioxidant system,enhancing the formation of organic acid-cadmium complexes with lower toxicity in the rhizosphere,and improving the stress resistance of seedlings to resist cadmium stress.更多还原