Comparative anatomy of embryogenic and non-embryogenic calli from Pimpinella brachycarpa

Anatomical differences between embryogenic and non-embryogenic calli of Pimpinella brachycarpa were investigated by light microscopy and electron microscopy. Initial callus tissue emerged from explants after 14 d of culturing. The embryogenic calli (EC) were firm, rather opaque, and light yellow in color. The cells usually formed small, compact clusters. Non-embryogenic calli (NEC), however, were friable, semitransparent, and yellow or gray. These formed relatively larger and loosely held clusters. Scanning electron microscopy showed that EC were composed of individual compact and spherical cells that were rather regular in size and approximately 20 μm long. All were tightly held together and appeared to organize globular embryos. In contrast, the NEC comprised elongated and loosely held cells that were approximately 50 μm long. Tubular and u-shaped NEC cells protruded irregularly, and were of varying heights along the cell aggregates. Transmission electron microscopy of the EC revealed typical eukaryotic cytoplasmic components, including nuclei, mitochondria, and vacuoles in the cytoplasm enclosed by an electron-transparent cell wall. Based on the numerous ribosomes within the cytoplasm, these cells appeared to be well-organized and metabolically active. The NEC cells were much larger and more highly vacuolated than those of the EC. In ultrathin sections, the former seemed to be almost devoid of other cellular contents except for plastids and nuclei. Furthermore, EC and NEC showed different regeneration capacities in their somatic embryo formation. Most EC produced hyperhydric somatic embryos, followed by normal somatic embryos; whereas only a few shooted or rooted somatic embryos arose from the NEC.