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Rhizomania is one of the most damaging and widely spread diseases in major sugar beet growing regions of the world. The causal agent, beet necrotic yellow vein virus (BNYVV), is transmitted via the fungus Polymyxa betae, which retains it in the field for years. In this study, an RNA silencing mechanism was employed to induce resistance against rhizomania using intron‐hairpin RNA (ihpRNA) constructs. These constructs were based on sequences of the BNYVV 5′‐untranslated region of RNA‐2 or the flanking sequence encoding P21 coat protein, with different lengths and orientations. Both transient and stable transformation methods produced effective resistance against rhizomania correlated with the transgene presence. Among the constructs, those generating ihpRNA structures with small intronic loops produced the highest frequencies of resistant events. The inheritance of transgenes and resistance was confirmed over generations in stably transformed plants.

Two diploid sugar beet genotypes of agronomical importance were transformed using Agrobactrium tumefaciens harboring pBI35Scry containing a synthetic cry1Ab gene. Leaf blade with attached shoot bases, a highly regenerative tissue, were used as explant substratum for transformation. PCR screening with cry1Ab-specific primers showed the presence of transgene in more than 50% of the regenerated kanamycin-resistant plants after treatment with the antibiotic. A transformation rate of 8.8-12.2% (depending on genotype) was achieved as revealed by genomic DNA dot blotting. The intact integration of transgene cassette into the genome was furthermore confirmed by Southern blot analysis. The expression of the cry1Ab gene encoding a truncated endotoxin (67 kDa) at about 0.1% of total soluble protein was achieved in the leaves of transgenic plants as shown by Western blot analysis. Bioassays under in vitro conditions with Spodoptera littoralis, one of the most important pests in sugar beet fields, demonstrated enhanced resistance against this pest. The inheritance of the inserted transgene was confirmed in F 1 plants obtained through crossing of T 0 plants with a cytoplasmic male sterile line. Transgenic plants are currently grown in a greenhouse and will be subjected to further bioassay analyses against other lepidopteran pests of sugar beet.

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Numerous diseases caused by fungal pathogens influence the annual production of sugar beet. In order to obtain a plant resistant to fungi, genetic transformation has been applied to the sugar beet. To invade a plant tissue, phytopathogenic fungi produce several cell wall-degrading enzymes (CWDEs); polygalacturonases (PGs) are pathogenicity factors produced at the earlier stages of a fungal infection that depolymerize the homogalacturonan. One of the strategies used by plants to limit the degradation of the cell wall polysaccharides by fungal CWDEs is the production of proteinaceous inhibitors. Against fungal, microbial, and insect PGs, plants produce cell wall-associated polygalacturonase-inhibiting proteins (PGIPs). The overexpression of PGIPs improves the resistance to fungal and bacterial necrotrophs in different plants. In this research, the gene encoding the PGIP1 fused downstream of the leader sequence for secretion in the extracellular environment was isolated from Phaseolus vulgaris and cloned into the expression vector pBI121 for the Agrobacterium-mediated transformation of sugar beet. Modified transformation protocol and selection strategies were developed. In comparison with the preexisting methods, the transformation efficiency was increased and different cultivars were transformed, highlighting the general effectiveness of the method applied. The presence of the transgene and the activity of PvPGIP1 were confirmed by PCR and agarose diffusion assay analyses, respectively, and the present and copy number of the transgene in the T0 plants' genome were demonstrated by Southern blot.

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