Convergence of Bar and Cry1Ac Mutant Genes in Soybean Confers Synergistic Resistance to Herbicide and Lepidopteran Insects.

Soybean is a globally important crop species, which is subject to pressure by insects and weeds causing severe substantially reduce yield and quality. Despite the success of transgenic soybean in terms of Bacillus thuringiensis (Bt) and herbicide tolerance, unforeseen mitigated performances have still been inspected due to climate changes that favor the emergence of insect resistance. Therefore, there is a need to develop a biotech soybean with elaborated gene stacking to improve insect and herbicide tolerance in the field. In this study, new gene stacking soybean events, such as bialaphos resistance (bar) and pesticidal crystal protein (cry)1Ac mutant 2 (M#2), are being developed in Vietnamese soybean under field condition. Five transgenic plants were extensively studied in the herbicide effects, gene expression patterns, and insect mortality across generations. The increase in the expression of the bar gene by 100% in the leaves of putative transgenic plants was a determinant of herbicide tolerance. In an insect bioassay, the cry1Ac-M#2 protein tested yielded higher than expected larval mortality (86%), reflecting larval weight gain and weight of leaf consumed were less in the T1 generation. Similarly, in the field tests, the expression of cry1Ac-M#2 in the transgenic soybean lines was relatively stable from T0 to T3 generations that corresponded to a large reduction in the rate of leaves and pods damage caused by Lamprosema indicata and Helicoverpa armigera. The transgenic lines converged two genes, producing a soybean phenotype that was resistant to herbicide and lepidopteran insects. Furthermore, the expression of cry1Ac-M#2 was dominant in the T1 generation leading to the exhibit of better phenotypic traits. These results underscored the great potential of combining bar and cry1Ac mutation genes in transgenic soybean as pursuant of ensuring resistance to herbicide and lepidopteran insects.

Verticillium dahliae transcription factors Som1 and Vta3 control microsclerotia formation and sequential steps of plant root penetration and colonisation to induce disease.

Verticillium dahliae nuclear transcription factors Som1 and Vta3 can rescue adhesion in a FLO8-deficient Saccharomyces cerevisiae strain. Som1 and Vta3 induce the expression of the yeast FLO1 and FLO11 genes encoding adhesins. Som1 and Vta3 are sequentially required for root penetration and colonisation of the plant host by V. dahliae. The SOM1 and VTA3 genes were deleted and their functions in fungus-induced plant pathogenesis were studied using genetic, cell biology, proteomic and plant pathogenicity experiments. Som1 supports fungal adhesion and root penetration and is required earlier than Vta3 in the colonisation of plant root surfaces and tomato plant infection. Som1 controls septa positioning and the size of vacuoles, and subsequently hyphal development including aerial hyphae formation and normal hyphal branching. Som1 and Vta3 control conidiation, microsclerotia formation, and antagonise in oxidative stress responses. The molecular function of Som1 is conserved between the plant pathogen V. dahliae and the opportunistic human pathogen Aspergillus fumigatus. Som1 controls genes for initial steps of plant root penetration, adhesion, oxidative stress response and VTA3 expression to allow subsequent root colonisation. Both Som1 and Vta3 regulate developmental genetic networks required for conidiation, microsclerotia formation and pathogenicity of V. dahliae.

A highly efficient Agrobacterium tumefaciens-mediated transformation system for the postharvest pathogen Penicillium digitatum using DsRed and GFP to visualize citrus host colonization.

Penicillium digitatum is a major postharvest pathogen of citrus crops. This fungus broadly spreads worldwide and causes green mold disease, which results in severe losses for citrus production. Understanding of the citrus infection by P. digitatum may help develop effective strategies for controlling this pathogen. In this study, we have characterized a virulent strain of P. digitatum isolated in Vietnam and established a highly efficient Agrobacterium tumefaciens-mediated transformation (ATMT) system for this fungal strain with two newly constructed binary vectors. These binary vectors harbor dominant selectable markers for hygromycin or nourseothricin resistance, and expression cassettes for the red fluorescent protein (DsRed) or the green fluorescent protein (GFP), respectively. Using the established ATMT system, the transformation efficiency of the Vietnamese strain could reach a very high yield of 1240±165 transformants per 106 spores. Interestingly, we found that GFP is much better than DsRed for in situ visualization of citrus fruit colonization by the fungus. Additionally, we showed that the transformation system can also be used to generate T-DNA insertion mutants for screening non-pathogenic or less virulent strains. Our work provides a new platform including a virulent tropical strain of P. digitatum, an optimized ATMT method and two newly constructed binary vectors for investigation of the postharvest pathogen. This platform will help develop strategies to dissect molecular mechanisms of host-pathogen interactions in more detail as well as to identify potential genes of pathogenicity by either insertional mutagenesis or gene disruption in this important pathogenic fungus.