Drought is one of the most common environmental factors that affect alfalfa germination and development. Nitric oxide (NO) could mediate stress tolerance in plants. The goal of this study was to determine exogenous NO donor–mediated drought adaption molecular mechanisms during the alfalfa germination stage. In this study, physiological and transcriptome analyses were performed on 7 days of the growth period seedlings by sodium nitroprusside (SNP) and polyethylene glycol (PEG) treatment. The results showed that SNP supplementation alleviated malondialdehyde accumulation, increased levels of proline and soluble sugars, and enhanced antioxidant enzyme activity under osmotic stress conditions. RNA-Seq experiments identified 5828 genes exhibiting differential expression in seedlings treated with PEG, SNP, or SNP+PEG relative to seedlings treated with distilled water. Of these DEGs, 3235 were upregulated, and 2593 were downregulated relative to the controls. Fifteen DEGs were amplified by qRT-PCR to verify the changes in expression determined by RNA-Seq, revealing that PIF3 , glnA , PLCG1 , and RP-S11e exhibited enhanced expression under the SNP+PEG treatment. SNP was found to modulate redox homeostasis-related genes such as GSTs , SOD2 , GPX , and RBOH , and triggered calcium signaling transduction. It also induced some key genes relating to the abscisic acid, ethylene, and auxin signaling transduction in response to PEG stress. Conversely, genes associated with secondary metabolite biosynthesis and the metabolism of starch and sucrose during osmotic stress were downregulated by SNP. These results provide new insights into SNP-mediated drought adaption mechanisms at transcriptome-wide in alfalfa and reveal key drought tolerance pathways in this species.

中文翻译：

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转录分析显示硝普钠影响苜蓿在萌发阶段对PEG诱导的渗透胁迫的反应

干旱是影响苜蓿发芽和发育的最常见的环境因素之一。一氧化氮 (NO) 可以介导植物的胁迫耐受性。本研究的目的是确定苜蓿萌发阶段外源 NO 供体介导的干旱适应分子机制。在这项研究中，通过硝普钠 (SNP) 和聚乙二醇 (PEG) 处理对生长期 7 天的幼苗进行生理和转录组分析。结果表明，在渗透胁迫条件下，补充 SNP 减轻了丙二醛的积累，增加了脯氨酸和可溶性糖的水平，并增强了抗氧化酶的活性。RNA-Seq 实验鉴定了 5828 个基因在用 PEG、SNP、或 SNP+PEG 相对于用蒸馏水处理的幼苗。在这些 DEG 中，相对于对照，3235 个被上调，2593 个被下调。通过 qRT-PCR 扩增了 15 个 DEG，以验证 RNA-Seq 确定的表达变化，揭示 PIF3、glnA、PLCG1 和 RP-S11e 在 SNP+PEG 处理下表现出增强的表达。发现 SNP 可调节氧化还原稳态相关基因，如 GST、SOD2、GPX 和 RBOH，并触发钙信号转导。它还诱导了一些与脱落酸、乙烯和生长素信号转导相关的关键基因，以响应 PEG 胁迫。相反，与次级代谢物生物合成以及渗透胁迫期间淀粉和蔗糖代谢相关的基因被 SNP 下调。