Somatic Embryogenesis of Immature Cunninghamia lanceolata (Lamb.) Hook Zygotic Embryos.

Two efficient somatic embryogenesis systems were developed in Chinese fir, the most important conifer for industrial wood production in China. Three development stages (cleavage polyembryony, dominant embryo, and precotyledon) of immature embryos derived from 25 genotypes of open-pollinated mother trees were used as initial explants. Cleavage polyembryony-stage embryos with a 12.44% induction rate was the most embryogenic response stage. The highest frequency of embryogenic callus (13.86%) induction was obtained from DCR medium supplemented with 1.5 mg L-1 2,4-dichlorophenoxyacetic acid (2,4-D) and 0.3 mg L-1 kinetin (KN). An average of 53.33 early somatic embryos were produced from approximately 0.2 g (fresh weight) embryogenic callus after 2 weeks of incubation on medium supplemented with 50 µmol L-1 abscisic acid (ABA) and 100 g L-1 polyethylene glycol (PEG) 6000. About 53% dominant embryos have an embryogenic response after a 6-week cultivation on medium supplemented with 1.0-2.0 mg L-1 benzyladenine (BA), 0.2 mg L-1 naphthylacetic acid (NAA) or 2,4-D, and 0.004 mg L-1 thidiazuron (TDZ). After three successive transfer cultures on medium supplemented with 1.5 mg L-1 BA, 0.2 mg L-1 NAA, and 0.004 mg L-1 TDZ, 4.49-16.51% of the embryos developed into somatic embryos.

Proteome profiling of early seed development in Cunninghamia lanceolata (Lamb.) Hook.

Knowledge of the proteome of the early gymnosperm embryo could provide important information for optimizing plant cloning procedures and for establishing platforms for research into plant development/regulation and in vitro transgenic studies. Compared with angiosperms, it is more difficult to induce somatic embryogenesis in gymnosperms; success in this endeavour could be increased, however, if proteomic information was available on the complex, dynamic, and multistage processes of gymnosperm embryogenesis in vivo. A proteomic analysis of Chinese fir seeds in six developmental stages was carried out during early embryogenesis. Proteins were extracted from seeds dissected from immature cones and separated by two-dimensional difference gel electrophoresis. Analysis with DeCyder 6.5 software revealed 136 spots that differed in kinetics of appearance. Analysis by liquid chromatography coupled to tandem mass spectrometry and MALDI-TOF mass spectrometry identified proteins represented by 71 of the spots. Functional annotation of these seed proteins revealed their involvement in programmed cell death and chromatin modification, indicating that the proteins may play a central role in determining the number of zygotic embryos generated and controlling embryo patterning and shape remodelling. The analysis also revealed other proteins involved in carbon metabolism, methionine metabolism, energy production, protein storage, synthesis and stabilization, disease/defence, the cytoskeleton, and embryo development. The comprehensive protein expression profiles generated by our study provide new insights into the complex developmental processes in the seeds of the Chinese fir.