Sorbitol and Sucrose-Induced Osmotic Stress on Growth of Wheat Callus and Plantlet Regeneration

Scutellar somatic embryos are most commonly used as a target tissue for transformation and regeneration of transgenic wheat plants. Tissue culture responsiveness of elite varieties of wheat is one of the critical factors limiting high-frequency transformation. This study investigated the influence of sorbitol and sucroseinduced osmotic stress on growth and plantlet regeneration in callus cultures of wheat (Triticum aestivum L. em Thell). Immature embryos, at early-medium milking stages, of field grown wheat cvs., HUW206, HUW234, Sonalika and HD2009, were induced to form the callus on MS medium supplemented with 2, 4-D and 3% sucrose alone/or in a combination of sucrose and sorbitol. Fresh weight of callus was significantly promoted when 1.5% (w/v) sorbitol was added to the MS medium in conjunction with 3% (w/v) sucrose. However, higher concentration of sorbitol concentration (3% w/v) in conjunction with 3% (w/v) sucrose did not further improve fresh weight. Sorbitol in combination with sucrose caused a decrease in the water content from calli and promoted embryogenic callus formation. After 4 weeks, a high frequency (10–25%) shoot formation was obtained on MS media, IAA 1 mgL¹ and zeatin 1 mgL⁻¹ in supplementation with sucrose and sorbitol. However, optimum plantlet regeneration (45–65%) was found on subsequent subculture on regeneration media containing 3% (w/v) sucrose and 3% (w/v) sorbitol. Osmotic stress induced by the combination of sucrose and sorbitol is beneficial for embryogenecity of the friable calli and high-frequency plantlet regeneration.