Cloning and Functional Analysis of ZFP5 from Amorpha fruticosa for Enhancing Drought and Saline–Alkali Resistance in Tobacco

Drought and soil salinization significantly constrain agricultural productivity, driving the need for molecular breeding strategies to enhance stress resistance. Zinc finger proteins play a critical role in plant response to abiotic stress. In this study, a gene encoding a C2H2-type zinc finger protein (<i>AfZFP5</i>) was cloned from <i>Amorpha fruticosa,</i> a species known for its strong adaptability. qRT-PCR analysis revealed that <i>AfZFP5</i> expression is regulated by sorbitol, H₂O₂, NaCl, and NaHCO₃. And all four treatments can cause upregulation of <i>AFZFP5</i> expression in the roots or leaves of <i>Amorpha fruticosa</i> within 48 h. Transgenic tobacco lines overexpressing AfZFP5 demonstrated enhanced tolerance to drought and salt-alkali stress at germination, seedling, and vegetative stages. Compared to wild-type plants, transgenic lines exhibited significantly higher germination rates, root lengths, and fresh weights when treated with sorbitol, NaCl, and NaHCO₃. Under natural drought and salt-alkali stress conditions, transgenic plants showed elevated activities of superoxide dismutase (SOD) and peroxidase (POD), and upregulated expression of oxidative stress-related kinase genes (NtSOD, NtPOD) during the vegetative stage. Additionally, transgenic tobacco displayed lower malondialdehyde (MDA) content and reduced staining levels with 3,3'diaminobenzidine (DAB) and Nitro blue tetrazolium (NBT), indicating enhanced reactive oxygen species (ROS) scavenging capacity by <i>AfZFP5</i> upon salt-alkali stress. Under simulated drought with PEG6000 and salt-alkali stress, chlorophyll fluorescence intensity and Fv/Fm values in transgenic tobacco were significantly higher than in wild-type plants during the vegetative stage, suggesting that <i>AfZFP5</i> mitigates stress-induced damage to the photosynthetic system. This study highlights the role of <i>AfZFP5</i> in conferring drought and salt-alkali stress tolerance, providing genetic resources and a theoretical foundation for breeding stress-resistance crops.