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

Abstract The cultivated diploid cotton species G. arboreum offers a better opportunity to elucidate gene structure and function compared to the allotetraploid cotton species through genetic transformation, the reliable and efficient method for high frequency somatic embryogenesis and plant regeneration in G. arboreum is urgent need to be established. Callus was induced from hypocotyl, root and cotyledon of G. arboreum seedlings on MSB (MS salts and B5 vitamins) medium with 0.09 µM 2,4-D and 2.32 µM KT. The embryogenic callus was induced on MS5 medium from the suspended cultures of several cycles of alternate liquid-solid culture, which was critical step for somatic embryogenesis. The liquid medium of MS4 was supplemented with 0.1g/L NaCl, 0.1g/L KCl and 0.1g/L CuSO4. The solid medium of MS5 for embryogenic callus effective induction was supplemented with 37.59 mM KNO3 + 62.47 µM NH4NO3 and 2.46 µM IBA + 0.93 µM KT or 0.045 µM 2,4-D + 2.46 µM IBA + 0.465 µM KT. During callus growing on different media, callus was effectively selected for subculture or treatment according to cell morphology to induce embryogenic callus and somatic embryos. Somatic embryo maturation and germination were better on MS5 medium with maltose or glucose + maltose than the single glucose. The regenerated plantlets with well-developed roots were directly transferred to soil or grafted onto the germinated cotton plantlets. The feasible process of plant regeneration via somatic embryogenesis in diploid cultivated species was established and needed to be improved and optimized for the gene functional analysis and gene editing in the diploid cotton species.