Comparison of Somatic Embryogenesis‐derived Coffee (Coffea arabica L.) Plantlets Regenerated in vitro or ex vitro: Morphological, Mineral and Water Characteristics

Abstract

Coffea arabica L. plantlets obtained ex vitro after sowing somatic embryos produced in a bioreactor in horticultural substrate were compared with those obtained in vitro from the same embryo population under conventional culturing conditions on semi-solid media. The intensity and quality of aerial and root system development were compared. Shoot emergence was more efficient in vitro but rooting frequencies were low. In contrast, all ex vitro-regenerated embryos rooted. The cotyledon area of mature embryos produced in a bioreactor positively affected plantlet development when regeneration was carried out ex vitro. Embryos with an intermediate cotyledon area (0.86 cm2) had the highest rates of plant conversion ex vitro (63%), and also resulted in vigorous plantlets. Mortality was higher in nursery conditions, but better plant development was obtained. The quality of plantlets produced under ex vitro conditions was reflected in better growth of the aerial and root systems, and also by similar morphological, mineral and water status characteristics to seedlings. Unlike roots formed on semi-solid media, those produced in soil were branched, fine (30-50% had a diameter of less than 0-5 mm) and they bore root hairs. Leaves of plantlets regenerated ex vitro had a histological structure similar to that of seedling leaves, and a lower stomatal density (100 vs. 233 mm-2). Moreover, they were more turgid, as indicated by higher pressure potential (psiP) (0.91 s. 0.30 MPa) and relative water content values (97 vs. 93%). Furthermore, under in vitro conditions, leaves had larger stomata which were abnormally round and raised. Direct sowing of germinated somatic embryos resulted in the rapid production of vigorous plantlets under ex vitro conditions, whilst removing the need for problematical and costly conventional acclimatization procedures.

TDZ pulsing evaluation on the in vitro morphogenesis of peach palm

“…2b-d), a phenomenon not reported to date and related to a low survival of adventitious roots during acclimatization (Smith and McClelland 1991;Barry-Etienne et al 2002).…”

Plant regeneration via indirect somatic embryogenesis and optimisation of genetic transformation in Coffea arabica L. cvs. Caturra and Catuaí

“…The regeneration medium R (van Boxtel and Berthouly, 1996) contained 17.6 µM of BAP whereas the S3 medium (Quiroz-Figueroa et al 2002) contained 2.32 µM of kinetin and 0.27 µM of napthaleneacetic acid (NAA). Somatic embryo development from embryogenic callus or suspension cultures has been accomplished previously in coffee mainly on semisolid culture media (Barry- Etienne et al 2002b;Barrueto et al 2004;Samson et al 2006). Temporary immersion culture systems have improved the development of somatic embryos in Citrus deliciosa, Hevea brasiliensis, Musa spp and C. arabica F1 hybrid (Caturra x E531) (Escalant et al 1994;Cabasson et al 1997;Etienne et al 1997;Barry-Etienne et al 1999;Barry-Etienne et al 2002a).…”

Bioreactors in coffee micropropagation

“…The direct sowing of pre-germinated embryos under ex vitro conditions, reduced handling times to 13 % and shelving area requirements to 6.3 % of the values obtained with conventional acclimatization of plants developed on semi-solid media and the time spent in vitro was reduced by 3 months (Barry- . Moreover, when compared to plantlets derived from the same embryo population obtained under conventional culturing conditions on semi-solid media, the plantlets obtained ex vitro after sowing somatic embryos were more vigorous (Barry- Etienne et al, 2002a).…”

“…ii) Neither is it obvious that bioreactors are essential for mass embryo production, since similar results in terms of quantity and quality to those obtained with conventional bioreactors have been obtained in Erlenmeyer flasks. However, embryos produced in temporary immersion bioreactors offer an advantage in terms of morphological quality (Barry- Etienne et al, , 2002a.…”

“…For Coffea canephora, Ducos et al (1999) proposed an efficient protocol for clone propagation in which the pre-germination phase was conducted on semi-solid medium in disposable Petri dishes. Similarly to the works of Barry- Etienne et al ( , 2002a, plant acclimatization and regeneration was achieved by directly sowing pre-germinated embryos under greenhouse conditions. That protocol, which does not envisage the use of bioreactors, is also being validated on a pre-industrial scale by that team (Ducos, personal comm.).…”