In-vitro Callogenesis in Safflower (Carthamus tinctorius L.) Genotypes from Leaf, Hypocotyl, and Floral Bud Explants under Differential Phytohormone Regimes

Abstract

Safflower (Carthamus tinctorius L.) is widely recognized for its pharmacologically active metabolites; however, its seasonal cultivation restricts consistent extraction of bioactive compounds. Plant tissue culture presents a sustainable strategy by enabling continuous and standardized metabolite production independent of field constraints. This study optimized callus induction from leaf, hypocotyl, and floral bud explants of two safflower genotypes, GMU-7931-1 and GMU-7923, using Murashige and Skoog (MS) medium supplemented with differential concentrations of NAA, BAP, 2,4-D, and TDZ. Marked explant and genotype specific responses were observed. In leaf explants, GMU-7931-1 exhibited the highest callus induction (86.66%) at 3.0 mg/L NAA + 1.0 mg/L BAP, while GMU-7923 responded maximally (90.00%) at 4.0 mg/L NAA + 1.0 mg/L BAP. Hypocotyl-derived callus was optimal at 2.0 mg/L NAA + 1.0 mg/L BAP for GMU-7931-1 (90%) and 3.0 mg/L NAA + 1.0 mg/L BAP for GMU-7923 (86.66%), whereas higher auxin levels (≥5.0 mg/L) suppressed callogenesis. For 2,4-D, peak responses were recorded at 5.0 mg/L in GMU-7931-1 (90%) and 3.0 mg/L in GMU-7923 (86.66%), with lower and higher levels resulting in reduced callus proliferation. Floral bud explants demonstrated robust callus formation and biomass accumulation on 0.5 mg/L NAA + 5.0 mg/L TDZ in both genotypes, with GMU-7923 showing superior performance. Notably, successful bud-derived callus induction reported here represents a novel addition to safflower tissue culture literature. Collectively, these findings establish remarkable response of  NAA (2.0–4.0 mg/L) with BAP (1.0 mg/L) for leaf and hypocotyl and 0.5 mg/L NAA + 5.0 mg/L TDZ for bud followed by GMU-7923 (10–12 day remarkable response at 3.0 mg/L 2,4-D). Across explants, leaf tissues consistently displayed superior callus induction, while GMU-7931-1 showed the earliest visible callus formation (8–10 days) under optimal 2,4-D treatments (5.0 mg/L) as highly effective regimes for efficient and reproducible callus induction, supporting their application in scalable metabolite and pigment production in Carthamus tinctorius.