Plant regeneration in Stone pine (Pinus pinea L.) by somatic embryogenesis

Abstract

Regeneration of plants by somatic embryogenesis (SE) was achieved in Stone pine (Pinus pinea), one of the most characteristic tree species of the Mediterranean ecosystem. The initial explants were megagametophytes containing zygotic embryos from five selected half-sib families collected at different dates over 2 consecutive years. Rates of extrusion and initiation of SE differed in both years. However, qualitative patterns were very similar: for most families, the responsive developmental window was from late cleavage polyembryony to early cotyledonary stage. The highest overall mean frequencies of extrusion and SE initiation (7 and 0.9%, respectively, for the five families and the eight 2006 collections) were obtained on a modified Litvay's medium with 9 lM 2,4-D and 4.5 lM BAP, supplemented with L-glutamine and casein hydrolysate. Families showed large differences in frequencies of SE initiation from year to year. Only seven embryogenic lines were induced in 2005, representing three of the five families tested, whereas 34 lines from all the families were obtained in 2006. Proliferation of embryonal masses (EM) was significantly improved when they were subcultured after dispersing in liquid medium and collected on filter paper disks, instead of being subcultured as small clumps. This effect showed a significant interaction with genotype. Several preconditioning treatments and culture media combinations were tested for embryo development and maturation. The high proliferation rate of EM hampered somatic embryo development. However, up to 42 mature embryos from different lines of three of the five families were obtained, 23 of them germinated and seven converted into somatic seedlings.

The effect of culture medium formulation on Pinus nigra somatic embryogenesis

“…In our experiments for Pinus nigra, by reducing the PGR content in the medium we obtained increase in the initiation frequencies for DCR medium in 2007 but experiments in the following years did not confirm the beneficial effect of lowered PGR concentration. For Pinus nigra, similarly as for other Pinus species (Becwar et al 1990, Miguel et al 2004, Klimaszewska et al 2001, Carneros et al 2009) the stage specific response of zygotic embryo explants to somatic embryogenesis initiation was observed and seemed to be an important factor. In our previous experiments we have used different concentration and combination of PGRs (2, 4-D, BA, NAA) for the initiation of somatic embryogenesis.…”

Bottlenecks in Pinus radiata somatic embryogenesis: improving maturation and germination

“…The initial amount of embryogenic tissue usually ranges from 500 mg (Maruyama et al 2007) to 50 mg (Carneros et al 2009) per filter paper disc (7 cm).…”

“…Although EM initiation protocols are now fairly well established, maturation of the EM into cotyledonary, normal somatic embryos is not always successful, in radiata pine and in other pine species. Problems such as low or asynchronous embryo production (Yildirim et al 2006;Breton et al 2005), abnormal morphology or poor root development, have been also reported for P. pinea (Carneros et al 2009) and P. kesiya (Choudhury et al 2008).…”

Inducción de callos embriogénicos y formación de proembriones somáticos en Pterogyne nitens Tull "tipa colorada"

“…Porcentaje de explantos en que ocurrió la formación de callos por tratamiento, en tres medios de cultivos, a los 45 días en Pterog yne nitens: A) hoja cotiledonar B) explantos foliares C) tallos. MSC= medio de MS con la fórmula original completa; MS50= medio al 50% de la concentración salina original y MSMOD= medio modificado solo en NH4NO3 y KNO3 medio (Carneros et al, 2009). En el presente trabajo, la reducción del contenido de nitrógeno en el medio (MS50 y MSMOD) no favoreció la respuesta callogé-nica de los explantos.…”

“…La embriogénesis somática ha sido reportada en varias especies, Pinus pinea (Carneros et al, 2009); Quercus rubra (Vengadesan y Pijut, 2009), Quercus suber ; Acrocomia aculeata (Ferreira Moura et al, 2008); Eucalyptus sp. (Pinto et al, 2008); Cupresssus sempervirens L. (Giovanelli y Carlo, 2007); Melia azedarach (Sharry et al, 2006); leguminosas (Marinucci et al, 2004); Araucaria angustifolia (Santos et al, 2002), Coffea canephora (López Gómez et al, 2010), Hevea brasiliensis (Lardet et al, 2009), Vitis sp.…”