De novo characterization of AcABI5 transcription factor and physiological responses to salt stress in Alhagi camelorum callus

Alhagi camelorum , a dominant leguminous shrub in the saline–hyperarid Taklimakan Desert, can complete its life cycle in salty soils, yet the molecular basis of seedling-stage salt tolerance remains unknown. Here, we developed an in-vitro callus system to dissect this trait without soil heterogeneity. Among 36 hormone regimes, 1.5 mg/L 2,4-D plus 0.5 mg/L 6-BA produced 100 % induction. Exposure of 28-d-old calli to 200 mM NaCl for 48 h caused transient swelling of cortical cells and a 5.3-fold rise in malondialdehyde (MDA). Antioxidant enzymes responded sequentially: superoxide dismutase (SOD) peaked at 6 h to scavenge superoxide, peroxidase (POD) maintained high activity throughout the first 24 h, and catalase (CAT) stabilized after 48 h, jointly keeping H₂O₂ below toxic levels. RNA-seq has identified an up-regulated transcription factor (log₂FC = 7) which was a basic leucine-zipper (bZIP) homologue of abscisic acid-insensitive protein 5 (ABI5). Quantitative RT-PCR confirmed 17-fold induction by NaCl and rapid decay after stress removal. Sub-cellular localization of a 35S::GFP-AcABI5 fusion in Nicotiana benthamiana epidermis showed exclusive nuclear fluorescence, consistent with a transcriptional regulator. Therefore, our study provides both optimized callus protocols and a candidate gene for engineering salt tolerance in A. camelorum and related desert legumes.