Genotype and auxin influence direct somatic embryogenesis from protoplasts derived from embryogenic cell suspensions of Asparagus officinalis L.

Abstract

Embryogenic callus from four asparagus genotypes, Jersey Giant No. 8, MD10, Rutgers 22, and 86SOM1 was simultaneously initiated from spear explants on semisolid LS medium containing 5 μM 2,4-D or 50 μM NAA. Calluses were used to initiate cell suspensions in liquid LS medium of the same composition. The eight sets of cell suspensions were used as protoplast donors at both 2 and 5 months of age. Protoplasts were immobilized at 10 5/ml density in MS medium with 0.6% agarose and overlaid with liquid KM medium; both containing the same type and concentration of auxin used for the corresponding donor cells or with plant growth regulator-free (PGR-free) medium. There was a significant interaction between genotype, suspension auxin, and inclusion or exclusion of PGRs in the protoplast culture media on plating efficiency, and colony and somatic embryo formation. Plating efficiencies at 14 days ranged from 0–40%. Globular somatic embryos developed directly from protoplasts in 10–14 days and bipolar embryos could be transferred in 3–4 weeks to embryo maturation medium (EM medium) composed of LS medium with 2% sucrose and 1% Phytagel. Conversion to plants occurred as rapidly as 1–2 weeks after transfer to EM medium or 5–6 weeks after initial protoplast culture. Although all four genotypes regenerated plants, Rutgers 22 had the highest conversion frequency at 42%. Most plants recovered from the 2,4-D-derived protoplasts were karyotypically aberrant while a higher frequency of normal plants were obtained from the NAA-derived protoplast cultures.