Microalgae and nanoparticles are currently considered promising tools for numerous agricultural and biotechnological applications. The green microalga Chlorella sp. MF1 and its biosynthesized silver nanoparticles (AgNPs) were used in this study as biofortification agents to enhance glucosinolate and kaempferol levels in Eruca sativa. UV–visible spectroscopy, XRD, FTIR and TEM were comprehensively used for characterizing Chlorella-based AgNPs. The biosynthesized AgNPs were found to be spherical in shape, with size ranging from 1.45 to 5.08 nm. According to FTIR measurements, silver ions were reduced to AgNPs by functional groups such as amide, hydroxyl and carboxylate. Different experimental treatments were conducted, including either soaking seeds of E. sativa or foliar spray with various concentrations of Chlorella suspension (1, 2, 3 and 4 g L−1) and AgNPs (5, 10, 20 and 40 mg L−1). Expression levels of five key genes in the biosynthetic pathway of glucosinolates (MAM1, SUR1, MYB34 and MYB51) and kaempferol (CHS) were assessed using qRT-PCR. The results indicated an upregulation in the gene expression levels in all treatments compared to control, recording the highest level at 40 mg L−1 AgNPs and 4 g L−1 Chlorella suspension. In addition, high glucosinolates and kaempferol content was detected in plants whose leaves were sprayed with AgNPs and Chlorella suspension (40 mg L−1 and 4 g L−1) based on HPLC analysis. Sequence analysis of amplified CHS fragments from E. sativa plants treated with AgNPs (40 mg L−1) showed high sequence similarity to A. thaliana CHS gene. However, there were several CHS regions with sequence polymorphism (SNPs and Indels) in foliar sprayed plants. Results of this study evidenced that the application of AgNPs and Chlorella suspension increased glucosinolates and kaempferol content in E. sativa through upregulation of key genes in their biosynthetic pathway.

中文翻译：

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银纳米粒子和小球藻处理诱导芥子中芥子油苷和山奈酚关键生物合成基因

微藻和纳米粒子目前被认为是许多农业和生物技术应用的有前途的工具。绿色微藻 Chlorella sp. 在本研究中，MF1 及其生物合成的银纳米粒子 (AgNPs) 被用作生物强化剂，以提高芥菜中的芥子油苷和山奈酚水平。紫外-可见光谱、XRD、FTIR 和 TEM 被综合用于表征基于小球藻的 AgNP。发现生物合成的 AgNPs 呈球形，尺寸范围为 1.45 至 5.08 nm。根据 FTIR 测量，银离子被酰胺、羟基和羧酸根等官能团还原为 AgNPs。进行了不同的实验处理，包括用不同浓度的小球藻悬浮液浸泡 E. sativa 种子或叶面喷洒 (1, 2, 3 和 4 g L-1) 和 AgNPs（5、10、20 和 40 mg L-1）。使用 qRT-PCR 评估了硫代葡萄糖苷（MAM1、SUR1、MYB34 和 MYB51）和山奈酚 (CHS) 生物合成途径中五个关键基因的表达水平。结果表明，与对照相比，所有处理的基因表达水平上调，在 40 mg L-1 AgNPs 和 4 g L-1 小球藻悬浮液中记录最高水平。此外，根据 HPLC 分析，在叶子喷洒 AgNPs 和小球藻悬浮液（40 mg L-1 和 4 g L-1）的植物中检测到高硫代葡萄糖苷和山奈酚含量。来自用 AgNPs (40 mg L-1) 处理的苜蓿植物的扩增 CHS 片段的序列分析显示出与拟南芥 CHS 基因的高度序列相似性。然而，叶面喷洒植物中有几个具有序列多态性（SNPs和Indels）的CHS区域。