Different cultivars of Vitis vinifera vary in their potential to form embryogenic tissues. The WUSCHEL (WUS)-related homeobox (WOX) genes have been shown to play an important role in coordinating the gene transcription involved in the early phases of embryogenesis. The expression dynamics of 12 VvWOX genes present in the V. vinifera genome in embryogenic and other tissues of 'Chardonnay' were analysed. In order to understand the influence of WOX genes on the somatic embryogenic process, their expression profiles were compared in two cultivars of V. vinifera ('Chardonnay' and 'Cabernet Sauvignon') that show different aptitudes for embryogenesis. The expression of all VvWOX genes was influenced by culture conditions. VvWOX2 and VvWOX9 were the principal WOX genes expressed during the somatic embryogenesis process, and the low aptitude for embryogenesis of 'Cabernet Sauvignon' was generally correlated with the low expression levels of these VvWOX genes. VvWOX3 and VvWOX11 were strongly activated in correspondence to torpedo and cotyledonary stages of somatic embryos, with low expression in the earlier developmental stages (pre-embryogenic masses and globular embryos) and during embryo germination. VvWOX genes appeared to be key regulators of somatic embryogenesis in grapevine, and the regulation of these genes during early phases of somatic embryogenesis differed between the two cultivars of the same species.

A novel method for initiating somatic embryogenesis in grapevine, based on immature whole flower culture, is presented. The embryogenic competence of flowers was compared to that of anthers and ovaries, the most widely used explant types, for five grapevine cultivars. Both the genotype and the explant source affected the differentiation of somatic embryos. The highest percentages of embryogenesis were obtained in ovary-derived calli from all cultivars tested with the exception of Brachetto a grappolo lungo. Whole flowers proved to be suitable material for initiating embryogenic cultures for most tested cultivars, and for 110 R, Chardonnay, and Grignolino they gave similar or better results than anthers. Collection of whole flowers from the inflorescence is easier and faster than excision of anthers and ovaries from the flower itself; it can be done without the use of a stereomicroscope and damage to the explant is unlikely. No morphological difference was noted among embryogenic cultures originated from ovaries, flowers, or anthers.

Indirect somatic embryogenesis is effective at eliminating the most important viruses affecting grapevines. Accordingly, this technique was tested as a method for eradicating two widespread viroids, Grapevine yellow speckle viroid 1 (GYSVd-1) and Hop stunt viroid (HSVd), from four grapevine cultivars. Both viroids were detected by RT-PCR in grapevine floral explants used for initiating embryogenic cultures, as well as in undifferentiated cells of embryogenic and non-embryogenic calli from anthers and ovaries. In contrast, somatic embryos differentiated from these infected calli were viroid-free, and viroids were not detected in embryo-derived plantlets even 3 years after their transfer to greenhouse conditions. A wider spatial distribution of HSVd than GYSVd-1 within proliferating calli was revealed by in situ hybridization, whereas no hybridization signal was detected in the somatic embryos. In addition, GYSVd-1 and HSVd were localised in the nucleus of infected cells, conclusively showing the nuclear accumulation of representative members of Apscaviroid and Hostuviroid genera, which has been only an assumption so far. Somatic embryogenesis was compared to in vitro thermotherapy, a technique routinely used for virus eradication. After thermotherapy, HSVd and GYSVd-1 were detected in all in vitro plantlets of the cultivar Roussan, and in all lines analysed after 3 years of culture in greenhouse. The high efficiency with which somatic embryogenesis may eliminate viroids and viruses from several infected grapevine cultivars, should allow the availability of virus-and viroid-free material, which would be useful not only for sanitary selection but also for basic research on plant-virus and plant-viroid interactions in grapevine.

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Floral biology of chestnut, from sporogenesis to mature embryo, is described. Microsporogenesis in flowers of unisexual catkins occurred in the first week of June 1991. Anthesis started in mid-June (≈70 days after budbreak) and lasted 2 weeks. In mid-June, in each pistillate flower, six to eight styles began to emerge, and 4 to 7 days later, they were extended fully (i.e., full bloom). In each flower, 10 to 16 anatropous ovules developed from the ovary axis. The megaspore mother cell had formed by the end of bloom. The mature ovule consisted of two integuments and a long, narrow nucellus with a small embryo sac of the Polygonum type. Zygotes were found 15 to 20 days after pistillate flower full bloom. Embryo development followed the Onagrad type, Trifolium variation. Seeds attained full size in mid-September, and fruit were mature in early November. The embryonal axis averaged 4.5 mm long × 2.1 mm wide. An apical meristem and the radicle were evident at opposite ends of the axis.