Plant regeneration via somatic embryogenesis in diploid cultivated cotton ( Gossypium arboreum L.)

The diploid cotton species G. arboreum offers a better opportunity to elucidate gene structure and function as opposed to the allotetraploid cotton species. As a prerequisite for this, a reliable and efficient method of high frequency plant regeneration in G. arboreum must be established. In this study, callus was induced from hypocotyl, root and cotyledon of G. arboreum seedlings on MSB medium (MS salts and B 5 vitamins) with 0.09 µM 2, 4-D (2, 4-dichlorophenoxyacetic acid) and 2.32 µM KT (kinetin). During propagation, callus was effectively selected for subculture on different media based on cell morphology to induce embryogenic callus and somatic embryos. Embryogenic callus was induced on MS 5 medium (MSB, 37.59 mM KNO 3 , 62.47 µM NH 4 NO 3 , 3% (w/v) glucose, 6.8 mM glutamine, 3.8 mM asparagine) from suspended cultures after several cycles of alternate solid MS 3 medium—liquid MS 4 medium culture, a key step in somatic embryogenesis. Solid MS 3 medium was supplemented with 2.46 µM IBA (īndole-3-butyric acid), 0.93 µM KT, 6.8 mM Gln, 3.8 mM Asn; liquid MS 4 medium was supplemented with 0.1 g/L NaCl, 0.1 g/L KCl and 0.1 g/L CuSO 4 . The solid–liquid alternate culture was effective for embryogenic callus induced in G. arboreum . MS 5 medium with maltose (3% w/v) or glucose (1.5% w/v) + maltose (1.5% w/v) performed better than that added single glucose (3% w/v) for somatic embryo maturation and germination. The regenerated plants with well-developed roots were directly transferred to the soil or grafted onto germinated cotton plants. Plant regeneration via somatic embryogenesis in diploid cultivated species was established, but there is a need for improvement and optimization for gene functional analysis and gene editing in the diploid cotton species.