Enhancement of somatic embryogenesis of Ipomoea batatas in solid eultures and production of mature somatic embryos in liquid cultures for application to a bioreactor production system

Abstract

Somatic embryos of Ipomoea batatas Lam. (sweet potato cv. 'White Star') were produced in an airlift bioreactor. This work describes the optimization of the embryogenic system on semisolid medium, followed by transfer of the system to liquid cultures and ultimately to the airlift bioreactor. The physiological age of embryogenic callus influenced the number and overall morphology of the embryo population in both semisolid and liquid medium. Maximum mature embryo production (35 embryos 10 mg-~ inoculum) was obtained from six-week-old callus at 30°C. Somatic embryogenesis occurred in liquid cultures containing 20 mM NH4NO3 and 30 mM KC1. Globular embryos formed and continued development in suspension producing viable, mature cotyledonary embryos by day 14. Embryo formation and development was limited in the bioreactor. Although shear stress was responsible for some embryogenic damage, the effect of purging the system with fresh air needed to be investigated. To isolate aeration effects from shear stress effects, atmospheric determinations were performed on shaker flask cultures. Initially the gas composition within the Erlenmeyer headspace was that of room air. Ethylene increased to a maximum of 6.4 ppm (day 16), maximum CO2, 21.2%, was also evident on day 16, and oxygen was depleted to a minimum of 8.1% by day 14. Purging the cultures with fresh air reduced the number of embryos formed; however, they were significantly longer than those formed in closed flasks. The gas response model of Ipomoea batatas will enable atmosphere replenishment in the bioreactor mimicking that of the shaker flask environment. Once the damaging effects of shear stress have been overcome, the regulation of bioreactor gasses should allow somatic embryo formation in the bioreactor comparable to that in shaker flasks.

Bioreactor technology for clonal propagation of plants and metabolite production

“…This can be manifested in the study on Taxus chinensis cell culture which revealed that both cell growth and production of taxuyunnanine C (TC) increased with increase in concentration of ethylene in bubble column bioreactor (Pan et al, 2000). However, Burg (1973) observed that ethylene inhibited cell division, and this might suppress embryo elongation (Bieniek et al, 1995).…”

“…For a well-functioning bioreactor, both the gas-liquid and liquid (medium)-solid (cells) mass transfers are important to conduct the respiration (metabolic process) of plant cells (Curtis, 2005). Carbon dioxide (CO 2 ), ethylene (C 2 H 4 ), ethanol (C 2 H 5 OH), and acetaldehyde (CH 3 CHO) are typically produced by respiring cultures (Thomas and Murashige, 1979;Perata and Alpi, 1991;Bieniek et al, 1995). Accumulation of these gases in the culture vessel inhibits growth of cultures (Jackson et al, 1985;Biddington and Robinson, 1991).…”

Effects from shear stress on morphology and growth of early stages of Norway spruce somatic embryos

“…The gas balance inside the bioreactor poses a challenge both in terms of obtaining a gas-balance permissible to somatic embryo development (Huang et al, 2006), as does managing the inhibitory shear stress from gas purging of the bioreactor (Bieniek et al, 1995). In our study, we have not studied the effect of the gas balance inside the flow cell.…”

In-vitro Techniques and Mutagenesis for the Improvement of Vegetatively Propagated Plants

“…Plant regeneration through organogenesis has been shown in roses, Rosa hybrida, Rosa chinensis minima (Hsia and Korban, 1996); mulberry, Morus alba (Susheelamma et al , 1996); papaya, Carica papaya (Drew, 1992); date palm, Phoenix dactylifera (Tisserat, 1984); and taro, Colocasia esculenta (Sabapathy and Nair, 1995). Somatic embryogenesis has been demonstrated in vegetables such as Allium jistulosum L. (Kim and Soh, 1996); potato, Solanum tuberosum (Garcia and Martinez, 1995); sweet potato, Ipomoea batatas (Bieniek et al, 1995;Sonnino and Mini, 1993); omamentals, such as roses, Rosa hybrida and Rosa chinensis minima (Hsia and Korban, 1996); oleander, Nerium oleander L. (Santos et al , 1994); cyclamen, Cyclamen persicum (Kreuger et al, 1995); gladiolus, Gladiolus hort. (Stefaniak, 1994); and fmit trees, such as banana, Musa sp.…”