In vitro regeneration of cotton (Gossypium hirsutum L.) cultivar KC3 with controlled phenolic secretion by using Kappaphycus alvarezii sulfated polysaccharide extract and plant growth regulators

Abstract

An efficient and reproducible in vitro regeneration protocol was developed for elite cotton cultivar KC3 by using plant growth regulators (PGRs) in combination with seaweed polysaccharide (SP) extracts. The existence of polysaccharide in seaweed extract was confirmed by Fourier transform infrared spectroscopy (FT-IR) and carbon-13 (¹³C) nuclear magnetic resonance (NMR) spectroscopy analysis. The extracted SP extract efficacy was tested for in vitro plant regeneration. The maximum callus frequency (89.4%) was obtained from hypocotyl explant in the Murashige and Skoog (MS) medium supplemented with 4.0% glucose, 1.5 mg L⁻¹ thidiazuron (TDZ), 0.6 mg L⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D), and 30.0 mg L⁻¹ SP. Remarkably, PGR- and SP-fortified medium inhibits the phenolic excretion from the explants. The well-developed yellow green friable texture of callus was transferred to shoot initiation medium. MS medium fortified with 4.0% glucose, 2.0 mg L⁻¹ 6-(γ,γ-dimethylallylamino) purine (2iP), 1.0 mg L⁻¹ kinetin (KIN), 1.0 mg L⁻¹ 6-benzylaminopurine (BA), and 40.0 mg L⁻¹ SP has shown maximum response (85.2%), and it produced 9.5 shoots per callus. The elongated shoots were cultured on root induction medium which consists of Murashige and Skoog (MS) salts with indole-3-acetic acid (IAA), indole-3-butyric acid (IBA), 1-naphthaleneacetic acid (NAA), and SP. The results revealed that the maximum number of roots (12.9 per shoot) with 8.6 cm in length was achieved on MS medium supplemented with 0.6 mg L⁻¹ IBA, combined with 30.0 mg L⁻¹ SP. Therefore, modified MS medium with natural bio-stimulant has more potential and is more reliable for in vitro regeneration of plants by neutralizing the effects of phenolic compounds secreted by the explants.