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褪黑素对盐胁迫下水稻种子萌发过程水分分布及种子活性的调控效应

Investigation on the Regulatory Effects of Melatonin on Internal Water Distribution and Seed Vigor During Rice Seed Germination Under Salt Stress

  • 摘要: 为探究褪黑素(melatonin,MT)对盐胁迫下水稻种子萌发过程中内部水分分布及种子活性的调控作用,该研究以粳型常规水稻“屉锦”为材料,通过NaCl浓度梯度筛选确定150 mmol/L(N150处理组)为标准盐胁迫浓度,进而设置清水对照组(CK1)、N150盐胁迫对照组(CK2)及5个MT浓度处理组(50、100、200、400、800 μmol/L MT+N150),结合低场核磁共振(low field nuclear magnetic resonance,LF-NMR)与核磁共振成像(magnetic resonance imaging,MRI)技术,对水稻种子萌发生理指标进行检测,探究MT对盐胁迫下水稻种子萌发的调控效应及水分代谢机制。结果表明:1)MT对盐胁迫下水稻种子萌发呈浓度依赖性双向调控,200 μmol/L为较优浓度,该浓度下种子发芽率、发芽势表现最佳,鲜重增长更接近CK1组,胚根、胚芽长度及次生根发育均优于其他处理组,可有效缓解盐胁迫引发的渗透压过高与离子毒害,维持细胞膜透性稳定;2)通过MRI伪彩映射处理可清晰呈现种子内部水分动态分布,结合生理代谢规律将萌发过程划分为吸胀期(0~24 h)、萌动期(24~48 h)、发芽期(48~72 h)3个阶段;3)将种子水分划分为强结合水(T20)、结合水(T21)、半结合水(T22)、自由水(T23)4种相态,其中中等浓度(200、400 μmol/L)的MT干预组表现出可维持水分相态稳定特性,其弛豫特征接近CK1组,而高浓度(800 μmol/L)组调控失效,水分相态特征趋近CK2组;4)综上,适宜浓度的MT处理能有效缓解盐胁迫对水稻种子萌发的抑制。利用LF-NMR及MRI技术可无损、精准解析种子内部水分动态与相态变化,该研究揭示了MT缓解盐胁迫下水稻种子萌发的内部水分变化规律,可为水稻种子耐盐性相关研究的开展及耐盐性提升技术的研发提供关键参考。

     

    Abstract: As a leading country in saline-alkali land distribution, China faces significant threats to its food security from soil salinization. Salt stress severely inhibits rice seed germination by inducing osmotic imbalance and ionic toxicity, presenting a major constraint for rice cultivation in these areas. Melatonin (MT), an indole derivative, has demonstrated efficacy in enhancing salt tolerance in various crops, primarily through reactive oxygen species (ROS) scavenging and ion homeostasis regulation. However, its specific regulatory mechanisms on internal water metabolism during the critical seed germination stage under salt stress remain poorly understood. The germination process is fundamentally driven by water uptake and redistribution, yet conventional methods lack the capability for real-time, non-destructive monitoring of internal water status and phase transitions. Low-field nuclear magnetic resonance (LF-NMR) and magnetic resonance imaging (MRI) techniques offer a powerful solution by enabling non-invasive detection of hydrogen proton signals, thereby precisely characterizing water content, migration, and phase distribution within biological tissues. This study investigated the regulatory effects of exogenous MT on germination characteristics and internal water dynamics in rice seeds under salt stress using the japonica conventional rice cultivar 'Tijin'. A preliminary NaCl concentration gradient screening established 150 mmol/L as the standard salt stress condition (N150 treatment), which reduced the germination potential to 32% of the water control. The experimental design included a water control group (CK1), a salt stress control group (CK2, N150), and five MT treatment groups (50, 100, 200, 400, and 800 μmol/L MT + N150). Germination physiological indices were monitored, and LF-NMR (utilizing CPMG sequence) combined with MRI (using MSE sequence) were employed to track internal water dynamics and phase changes throughout the 72-hour germination period. The results revealed a concentration-dependent biphasic regulation of rice seed germination by MT under salt stress. The 200 μmol/L MT concentration was identified as optimal, significantly enhancing germination rate (94.33%), germination potential (78.67%), fresh weight accumulation, and the growth of radicles, plumules, and secondary roots. This treatment effectively alleviated salt-induced osmotic stress and ionic toxicity, maintaining stable cell membrane permeability. MRI pseudo-color mapping vividly illustrated the dynamic internal water distribution, allowing the germination process to be delineated into three distinct stages based on physiological metabolism: imbibition (0-24 h), initiation (24-48 h), and germination (48-72 h). LF-NMR T2 relaxation analysis characterized four water fractions: strongly bound water (T20, 0.1-1 ms), bound water (T21, 1-10 ms), semi-bound water (T22, 10-100 ms), and free water (T23, 100-1000 ms). Medium MT concentrations (200 and 400 μmol/L) effectively maintained water phase stability, exhibiting relaxation characteristics akin to the CK1 group. In contrast, the high MT concentration (800 μmol/L) failed to confer beneficial effects, with its water phase features, including a consistently reduced T22 peak area, converging towards those of the salt-stressed CK2 group. In conclusion, an appropriate concentration of exogenous MT (200 μmol/L) can effectively mitigate the inhibitory effects of salt stress on rice seed germination. The integration of LF-NMR and MRI techniques provides a robust, non-destructive, and precise methodology for analyzing internal water dynamics and phase transitions. This study elucidates the patterns of internal water changes associated with MT-mediated alleviation of salt stress during rice seed germination, offering valuable theoretical insights and a technical reference for advancing research on rice seed salt tolerance and the development of related enhancement strategies.

     

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