Transgenic rice (Oryza sativa L.) lines were generated through particle-bombardment-mediated transformation. Hygromycin phosphotransferase (hpt) was used as a selectable marker gene on co-integrate plasmids containing either one or two unselected genes, the Bialaphos-resistance gene (bar) coding for phosphinothricin acetyltransferase and the b-glucuronidase gene (gusA), respectively. Transformants were analyzed to determine possible correlation between expression, integrated transgene copy number and/or complexity of integration patterns. We observed that an increase in transgene copy number did not always lead to a concomitant decrease in expression levels or to silencing through co-suppression. Transgenic lines with four to ®ve copies of integrated transgenes expressed the protein product of both unselected genes stably and at levels comparable to transformants with one or two copies. In the majority of lines we analyzed, expression patterns of the two unselected genes were similar. In lines where transgene silencing was observed, this was independent of position eects. In speci®c cases, silenced transgenes could be reactivated by treatment with 5-azacytidine, suggesting methylation of cytosine residues. We report that methylation of cytosines may not spread to adjacent regions, hence other transgenes in the vicinity of the silenced transgene remain active. By comparing the structure of transgenic loci with expression patterns of introduced genes, we concluded that the integrity of integrated transgenes was a major factor in the onset of silencing. We observed that the presence of truncated sequences of transgenes capable of generating incomplete transcripts, resulting in aberrant RNA species, may be responsible for silencing.

Bacillus thuringiensis (Bt) crystal protein genes encode insecticidal ␦-endotoxins that are widely used for the development of insectresistant crops. In this article, we describe an alternative transgenic strategy that has the potential to generate broader and more sustainable levels of resistance against insect pests. Our strategy involves engineering plants with a fusion protein combining the ␦-endotoxin Cry1Ac with the galactose-binding domain of the nontoxic ricin B-chain (RB). This fusion, designated BtRB, provides the toxin with additional, binding domains, thus increasing the potential number of interactions at the molecular level in target insects. Transgenic rice and maize plants engineered to express the fusion protein were significantly more toxic in insect bioassays than those containing the Bt gene alone. They were also resistant to a wider range of insects, including important pests that are not normally susceptible to Bt toxins. The potential impact of fusion genes such as BtRB in terms of crop improvement, resistance sustainability, and biosafety is discussed.Bt genes ͉ transgenic plants ͉ transgenic maize ͉ transgenic rice T ransgenic plants expressing Bacillus thuringiensis (Bt) toxins have been used successfully to provide resistance against selected insect pests for several years. Indeed, insect resistance is the second most widely used trait in transgenic crops (after herbicide tolerance) in world agriculture (1-5). One potential problem with Bt genes is that Bt insecticides are very widely used, with up to 90% of microbiological insect control products based on topically applied Bt toxins. For this reason, there is concern that insects might evolve resistance to Bt toxins (6, 7). The diamondback moth (Plutella xylostella) has evolved resistance in some open field populations in response to repeated exposure to foliar sprays containing Bt proteins (8), whereas laboratory selection experiments with other insect pests have shown that recessive mutant alleles can confer resistance to multiple Bt toxins (7-10). However, note that the evolution of resistance in insects against transgenic plants expressing Bt toxins has yet to be seen in the field.Recent strategies to address potential limitations in conventional transgenic insect pest control include the stacking or pyramiding of multiple transgenes in the same transgenic plant (11) and the use of hybrid toxins (e.g., fusions between a synthetic truncated Cry1Ba and domain II from Cry1Ia; ref. 12). We have devised an alternative strategy in which the Bt toxin Cry1Ac is fused to the nontoxic ricin B-chain (RB). The recognition of toxin-binding sites in the insect midgut is an important factor determining the spectrum of Bt toxin activity and the severity of toxemia (13). Several groups investigating the mechanism of toxin recognition have identified N-acetyl galactosamine residues as an important component of Bt toxinbinding receptors (14-16). Therefore, we selected the ricin B subunit as a fusion partner for the Bt toxin because RB is a galactose...

Knowledge on gene action and trait expression are important for effective breeding. The objective of this study was to determine the general combining ability (GCA), specific combining ability (SCA), maternal effects and heritability of drought tolerance, yield and yield components of candidate sweetpotato clones. Twelve genotypes selected for their high yield, dry matter content or drought tolerance were crossed using a full diallel mating design. Families were field evaluated at Masoro, Karama, and Rubona Research Stations of Rwanda Agriculture Board. Success rate of crosses varied from 1.8 to 62.5% with a mean of 18.8%. Family by site interaction had significant effect (P < 0.01) on storage root and vine yields, total biomass and dry matter content of storage roots. The family effects were significant (P < 0.01) for all parameters measured. Broad sense heritability estimates were 0.95, 0.84, 0.68, 0.47, 0.74, 0.75, 0.50, and 0.58 for canopy temperature (CT), canopy wilting (CW), root yield, skin color, flesh color, dry matter content, vine yield and total biomass, respectively. The GCA effects of parents and SCA effects of crosses were significant (P < 0.01) for CT, CW, storage root, vine and biomass yields, and dry matter content of storage root. The ratio of GCA/SCA effects for CT, CW, yield of storage roots and dry matter content of storage roots were higher than 50%, suggesting the preponderance of additive over non-additive gene action in the expression of these traits. Maternal effects were significant (P < 0.05) among families for CT, CW, flesh color and dry matter content, vine yield and total biomass. Across sites, the best five selected families with significant SCA effects for storage root yield were, Nsasagatebo × Otada 24, Otada 24 × Ukerewe, 4-160 × Nsasagatebo, K513261 × 2005-034 and Ukerewe × K513261 with 11.0, 9.7, 9.3, 9.2, 8.6 t/ha, respectively. The selected families are valuable genetic resources for sweetpotato breeding for drought tolerance, yield and yield components in Rwanda or similar environments.

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