Transformation of sugar beet with a bean chitinase gene and enhanced resistance to Alternaria alternata

Abstract

Fungal diseases are the major factors of sugar beet yield losses worldwide. Expression of pathogenesis-related proteins such as chitinases is considered as one of the plant defense responses against pathogens. Chitinases are cell wall degrading enzymes which have been shown to have high antifungal activity against a wide range of phytopathogenic fungi. In this study, two diploid sugar beet genotypes, SBSI-02 and SBSI-04, were used for transformation through Agrobacterium tumefaciens strain LBA4404 harboring the pBI-BCH plasmid containing the chi gene under the control of the CaMV35S promoter and the nptII selectable marker gene. Leaf blade with attached shoot bases were used as explant substratum for transformation. Green shoots were successfully screened using different concentration of kanamycin. PCR screening with chi-specific primer showed the presence of the transgene in more than 50% of regenerated kanamycin-resistant plants. Dot blot analysis confirmed the integration of at least one copy of the chi gene into the genome of putative transgenic plants. Western blot analysis revealed chitinase protein accumulation in transgenic plants. The content of chitinase protein varied among the five T0 transgenic plants. Bioassay analysis using detached leaves and at the whole plant level revealed increased resistance of T0 transgenic sugar beet lines against the fungal pathogen Alternaria alternata compared to non-transformed control plants.

Keywords

• sugar beet (beta vulgaris l.)
• agrobacterium tumefaciens
• chitinase gene
• transformation
• alternaria alternata