Variations in Somatic Polyembryogenesis: Induction of Adventitious Embryonal-Suspensor Masses on Developing Douglas Fir Embryos

Abstract

Embryonal-suspensor masses (ESMs) from seven commercial genotypes of Douglas fir are distinguished from embryogenic calluses by origin, cytochemical staining and developmental sequence. Zygotic ESMs with a dominant embryo can be rescued and new embryos multiplied and reconstituted in darkness by cleavage and budding polyembryony without a callus phase. New ESMs can also be induced directly on the surface of developing embryos. By contrast, the recovery of embryos derived from an intermediary callus phase follows a spatio-temporal wave of activity from the radicle towards the cotyledons starting 4 to 8 weeks after fertilization. In all embryo recovery systems, callus formation and teratogenic structures may be avoided by careful subculture, minimal exposure to low levels of plant hormones, and by selection of true-to-type ESMs grown in darkness.

Callusing of protodermal cells is promoted in light. The dedifferentiated cells can be induced to redifferentiate back into embryogenic ESMs in darkness. Callus-based regenerative processes can be followed microscopically by the increased formation of acetocarmine-reactive factors produced by nuclei and amyloplasts of redifferentiating cells. These factors are absent in nonembryogenic calluses. The alternative origins of ESMs are important to distinguish because the mechanisms by which proembryos are reconstituted and multiplied may contribute to different genotypic and phenotypic variances in regenerated products.

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