Embryogenic callus of sweetpotato [Ipomoea batatas (L.) Lam.] disassociates in liquid medium to form a heterogeneous population of embryogenic and nonembryogenic cell aggregates of varying sizes. To improve embryo production, such cell aggregate populations were obtained by manually fragmenting calli 5 to 10 mm in size into liquid medium. The resulting suspensions were analyzed and the embryogenic fraction identified. The percentage of embryogenic aggregates and the percentage of aggregates forming embryos decreased with decreasing aggregate size. Thus, 76% of the 710- to 1000-μm-diameter aggregates but only 14% of the 180- to 250-μm aggregates had embryogenic potential. However, only 20% of the 710- to 1000-μm aggregates and only 2% of the 180- to 250-μm aggregates actually formed embryos. Conversely, embryogenic callus and embryo production per milligram of cultured embryogenic callus increased quadratically with decreasing aggregate size. Individual torpedo-stage embryos were produced from cell aggregates 180 to 250 μm in size.
Raymond P. Chée and Daniel J. Cantliffe
Raymond P. Chée, Daniel I. Leskovar and Daniel J. Cantliffe
Embryogenic callus growth of sweetpotato [Ipomoea batatas (L.) Lam.] was selectively enhanced by subculture on basal callus proliferation medium modified to contain 15 mm NH4NO3. Embryogenic callus production was doubled on basal callus proliferation medium modified to contain 60 mm K+, while nonembryogenic callus production was reduced 40%. Additions of up to 40 mm NaCl to basal callus proliferation medium did not affect callus proliferation. The development of embryos from calli subculture to embryo production basal medium was unaffected by the KCl or NaCl treatments of the callus proliferation phase. However, embryo production was increased by subculturing callus from callus proliferation medium containing 20 mm NH4 + to embryo production medium containing 10 mm NH4 + Our results demonstrate that changes in mineral nutrition, in addition to growth regulator differences between callus proliferation and embryo production media, are important factors in sweetpotato somatic embryogenesis.
Yehoshua Saranga, David Rhodes and Jules Janick
Tolerance to partial desiccation and amino acid composition of celery (Apium graveolens L. cv. SB 12) somatic embryos were investigated under various culture durations and with exogenous application of 1 μm ABA, proline, and/or γ -aminobutyrate (GABA). ABA consistently increased tolerance to partial desiccation and elevated proline and GABA content of embryos. The changes in tolerance to partial desiccation associated with changes in culture duration (optimum 9 to 10 days) correlated with embryo proline content. Exogenous proline increased embryo proline content and tolerance to partial desiccation. Exogenous GABA increased embryo GABA content and tolerance to partial desiccation only when applied in combination with proline. Chemical name used: abscisic acid (ABA).
Raymond P. Chée, Jonathan R. Schultheis and Daniel J. Cantliffe
Plant formation from somatic embryos in response to BAP, NAA, and sucrose was studied in sweetpotato [Ipomoea batatas (L.) Lam.]. A maximum of 15% embryos at the torpedo stage of development formed plants of agar-solidified basal medium containing 3% sucrose and no growth regulators. The percentage of embryos forming shoots was increased to 53% by 4 μm BAP, but BAP reduced whole plant formation and promoted callusing at the root axis end of embryos. The frequency of plant development was increased to 38% by adding 0.1 μm NAA to the basal medium. Reducing sucrose concentration to 1.6% in basal medium increased the frequency of plant development to 32%. Chemical names used: 6-benzylaminopurine (BAP; α-naphthaleneacetic acid (NAA).
Jonathan R. Schultheis, Daniel J. Cantliffe and Herbert H. Bryan
Early plant growth, root quality, and yield from sweetpotato plants obtained from zygotic seed, somatic embryos, or cloned from stock plants (through micropropagation, rooted node explants, or nonrooted terminal vine cuttings) were compared in field plantings established in 1986, 1987, and 1988 in Gainesville and/or Homestead, Fla. At planting, transplants derived from somatic embryos had more nodes than the other propagules, while vine length per plant was greatest with nonrooted vine cuttings obtained from stock plants. The number of nodes (up to 253%) and vine growth (up to 517%) were greater when plants were derived from stock plants and zygotic embryos than from somatic embryos 4 weeks (1987) and 6 weeks (1988) after planting. Vegetative growth, larger-sized storage roots (>6 cm in diameter), and total yields (all root grades combined) were consistently reduced when plants were derived from somatic embryos compared with propagules of stock plant origin. Plants obtained from somatic embryos required more time for roots to bulk or size than the other propagule types. Root yield from plantlets derived from somatic embryos showed a 14-fold increase when harvest was delayed at least 53 more days. Root weight, regardless of harvest date, was greater when plants were derived from stock plants rather than from somatic embryos, while in most cases plants derived from somatic embryos yielded a greater number of roots than from stock plants. Plants obtained through somatic embryony and harvested at a later date typically had yields exceed 1.8 kg per plant. Morphology of plants obtained from somatic embryos was uniform and identical to plants derived from stock plants.
Yehoshua Saranga, Y.-H. Kim and Jules Janick
Effects of reduced osmotic potential on somatic embryos of celery (Apium graveolens L.) were studied in an attempt to understand and improve their tolerance to partial desiccation. Embryos responded similarly to application of high osmoticum (384 mOs/kg H2O vs. 190 mOs/kg H2O in the control), achieved either by manipulation of sucrose or polyethylene glycol concentrations (PEG). Treatments of high osmotic concentration applied during the last 2 days of the embryo production cycle increased embryo survival and conversion after partial desiccation. The most striking effect of the high osmotic concentrations was the 4-fold increase in proline, while a 2-fold increase was obtained with 1 μm ABA alone. Application of high osmotica decreased reducing sugars, increased sucrose, but did not affect starch content of embryos; of these responses, only the change in sucrose was similar to that induced by ABA. Osmotic treatments did not affect total fatty acid content in the embryos compared to the 2-fold increase induced by ABA. Chemical name used: abscisic acid (ABA).
Nirmal Joshee, Bipul K. Biswas and Anand K. Yadav
as leaf explants followed by development of successful and viable synthetic seeds. We also provide information on the encapsulation of somatic embryos for conservation and low-temperature storability of germplasm. Materials and Methods
Yehoshua Saranga, David Rhodes and Jules Janick
Desiccation tolerance of somatic embryos is a key factor for production of dry synthetic seeds. In celery (Apium graveolens L.) desiccation tolerance can be enhanced by optimization of culture duration, ABA application, or sucrose concentration in the embryo production medium. Morphologically mature embryos cultured for 10 days have shown higher desiccation tolerance then those cultured for 8 days, indicating that biochemical changes occur without any noticeable morphological changes. Application of ABA (1 μM) for the last two days of the embryo production cycle was critical for inducing desiccation tolerance; ABA application for the last four days had some additional beneficial effect. Desiccation tolerance was further enhanced by increasing the sucrose concentration of the embryo production media from 3% to 7% for the last two days. Increased desiccation tolerance achieved with optimal harvest timing and ABA application were associated with increased endogenous proline and aminobutyrate, and reduced glutamine.
Traud Winkelmann, Lara Meyer and Margrethe Serek
Somatic embryos of cyclamen [Cyclamen persicum Mill.] were produced using a liquid culture system. Two encapsulation techniques, conventional alginate beads and alginate hollow beads, were tested for globular cyclamen somatic embryos with the aim of developing synthetic seeds. Final germination from alginate beads was as high as observed for non encapsulated control embryos (97%), but germination was delayed. In contrast, germination from hollow beads was lower (71%) and occurred later. In hollow beads somatic embryos developed within the capsule, and outgrowth seemed to be more difficult than from alginate. Storage at 4 °C for four weeks resulted in a reduction of viability for controls as well as for encapsulated embryos. Incorporation of medium into the capsules improved the speed of germination for both capsule types. However, somatic embryos were not able to germinate on a medium-free support, even if encapsulated in beads containing medium.
Kalavathy Padmanabhan, Daniel J. Cantliffe, Roy C. Harrell and Dennis B. McConnell
A comparison of external morphology captured via a computer vision system and a study of internal anatomy of sweetpotato somatic embryos identified five different major morphological variants among torpedo and cotyledonary stage embryos. These included 1) Perfect Type, 2) Near Perfect Type, 3) Limited/No Meristematic Activity Type, 4) Disrupted Internal Anatomy Type, 5) Proliferating Type. Perfect and Near Perfect types of somatic embryos were categorized as competent, while Limited/No Meristematic activity, Disrupted Internal Anatomy, and Proliferating types were categorized as noncompetent with respect to their conversion ability. Lack of organized shoot development in somatic embryos of sweetpotato was attributed to the following abnormalities: 1) lack of an organized apical meristem, 2) sparsity of dividing cells in the apical region, 3) flattened apical meristem, 4) multiple meristemoids and/or diffuse meristematic activity throughout the embryo. A morphological fate map of most of the torpedo and cotyledonary embryo variants was identified, which will be beneficial in synthetic seeding and transgenic research and development of sweetpotato.