SummaryTransgenic plants are attractive biological systems for the large-scale production of pharmaceutical proteins. In particular, seeds offer special advantages, such as ease of handling and long-term stable storage. Nevertheless, most of the studies of the expression of antibodies in plants have been performed in leaves. We report the expression of a In addition, we demonstrate that a plant-made antibody with triantennary highmannose-type N -glycans has similar Fab functionality to its counterpart with biantennary complex N -glycans, but the former antibody interacts with protein A in a stronger manner and is more immunogenic than the latter. Such differences could be related to a variable immunoglobulin G (IgG)-Fc folding that would depend on the size of the N -glycan.

The promoter from rice tungro bacilliform badnavirus (RTBV) is expressed only in phloem tissues in transgenic rice plants. RF2a, a b-Zip protein from rice, is known to bind to the Box II cis element near the TATA box of the promoter. Here, we report that the full-length RTBV promoter and a truncated fragment E of the promoter, comprising nucleotides ؊164 to ؉45, result in phloemspecific expression of ␤-glucuronidase (GUS) reporter genes in transgenic tobacco plants. When a fusion gene comprising the cauliflower mosaic virus 35S promoter and RF2a cDNA was coexpressed with the GUS reporter genes, GUS activity was increased by 2-20-fold. The increase in GUS activity was positively correlated with the amount of RF2a, and the expression pattern of the RTBV promoter was altered from phloem-specific to constitutive. Constitutive expression of RF2a did not induce morphological changes in the transgenic plants. In contrast, constitutive overexpression of the b-ZIP domain of RF2a had a strong effect on the development of transgenic plants. These studies suggest that expression of the b-Zip domain can interfere with the function of homologues of RF2a that regulate development of tobacco plants. R egulation of transcription is achieved by the activity of multiple proteins that bind to regulatory elements, many of which are upstream of the promoters and alter basal rates of transcription initiation and͞or elongation (1, 2). To understand the mechanisms of tissue-specific and constitutive gene expression in plants, a number of promoters and transcription factors have been studied in recent years (3-16). It was shown that constitutive promoters, such as the cauliflower mosaic virus 35S promoter (17) and the promoter from cassava vein mosaic virus (14) are modular in organization and multiple cis elements. These elements, with specific transcription factors, apparently interact in an additive and͞or synergistic manner to confer gene expression in all plant tissues. Similarly, tissue-specific promoters contain multiple elements that contribute to promoter activity in both positive and negative ways (4-6, 8, 12, 18).The rice tungro bacilliform badnavirus (RTBV) promoter (19,20) and the transcription factors that interact it may serve as a model system to study plant tissue-specific gene expression. The virus, which replicates solely in phloem tissues, has a single promoter that is active in transfected protoplasts and is phloemspecific in transgenic rice plants (7,10,21,22).Within the fragment E of the promoter (nucleotides Ϫ164 to ϩ45), multiple cis elements were identified as being required for phloem-specific gene expression (7,10,15,23). A b-ZIP type transcription factor, RF2a, was isolated from rice and bound to Box II, a crucial cis element of the promoter. RF2a activates transcription from the RTBV promoter in an in vitro transcription system derived from rice cell cultures (11). Moreover, studies in transgenic rice plants suggested that RF2a is involved in the development of vascular tissues (11).We report here the functi...

A protocol was established for the introduction of DNA into embryogenic suspension-derived tissues of cassava via microparticle bombardment, for the selection of genetically transformed cells, and for the regeneration of fully transgenic plants from these cells. The plasmid DNA used for bombardment contained a gene encoding neomycin phosphotransferase (nptII) and a gene encoding beta-glucuronidase (uidA). Selection of bombarded tissue with paromomycin resulted in the establishment of putative transgenic embryogenic calli. In most of these calli, beta-glucuronidase was detected histochemically. Molecular analysis of paromomycin-resistant embryogenic calli and of plants regenerated from these calli, confirmed the stable integration of bombarded DNA into the cassava genome.

Methods for improving the efficiency of plant recovery from somatic embryos of cassava (Manihot esculenta Crantz) were investigated by optimizing the maturation regime and incorporating a desiccation stage prior to inducing germination. Somatic embryos were induced from young leaf lobes of in vitro grown shoots of cassava on Murashige and Skoog medium with 2,4-dichlorophenoxy acetic acid. After 15 to 20 days of culture on induction medium, the somatic embryos were transferred to a hormone free medium supplemented with activated charcoal. In another 18 days mature somatic embryos became distinctly bipolar and easily separable as individual units and were cultured on half MS medium for further development. Subsequent desiccation of bipolar somatic embryos resulted in 92% germination and 83% complete plant regeneration. The plants were characterized by synchronized development of shoot and root axes. Of the non-desiccated somatic embryos, only 10% germinated and 2% regenerated plants. Starting from leaf lobes, transplantable plantlets were derived from primary somatic embryos within 70 to 80 days.

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The complete sequence of 8159 nucleotides of the double stranded DNA genome of cassava vein mosaic virus (CsVMV) was determined (# U59751) and revealed a significant difference in genome organization when compared with a previous report (# U20341). When transferred to cassava plants by microbombardment, the full length CsVMV clone was infectious, confirming the genome organization here described. Sequence comparisons between CsVMV and members of the genera Caulimovirus and Badnavirus revealed high homologies between consensus sequences of several proteins that are indispensable for virus replication, including a potential transactivator factor not reported previously. The presence of a sequence complementary to a plant Met tRNA confirms that CsVMV is a plant pararetrovirus and is most closely related to members of the genus Caulimovirus as previously assessed. However, differences in genome organization, number and size of the ORFs, in addition to sequence comparisons with other plant pararetroviruses, shows that either the genetic variability of caulimoviruses is much greater than previously thought, or that CsVMV is the unique representative of a new genus within the Caulimoviridae family. On the basis of this study, it is proposed to upgrade the floating genus Caulimovirus to the family level and to divide the Caulimoviridae family into at least three genera with CsVMV being the type member of a new genus.

Nodes from 3-to 5-week-old in vitro plants of different cassava cultivars were cultured for 2-3 days on solid Murashige and Skoog basal medium supplemented with cytokinin to induce the enlargement of axillary buds. Subculture of these buds on the same medium resulted in multiple shoot formation within 4-6 weeks. Of the four cytokinins tested (6-benzylaminopurine (BAP), thidiazuron (TDZ), zeatin, and kinetin), BAP induced shoot development most efficiently. The best results were obtained with cultivar TMS 30555, in which 63% of the explants each produced at least 25 shoots on medium with 10 rag/1 BAR In cultivars that did not produce shoots, the addition of the surfactant Pluronic F-68 (2% wt/vol) raised the percentage of explants forming at least 5 shoots from 0 to 20-60%. Axillary buds were also used to dissect meristems and test their ability to regenerate into shoots. Shoot formation from meristems of six different cultivars was observed after preculture on medium with 5 mg/1 BAP followed by transfer to 10 mg/1 BAR

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