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KMID : 0903619970380020188
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Journal of the Korean Society for Horticultural Science
1997 Volume.38 No. 2 p.188 ~ p.193
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Transformation of Gladiolus for Disease Resistance
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Kamo, K.
Hammond, J./Roh, M.
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Abstract
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Transgenic Gladiolus plants containing the gusA gene under the cauliflower mosaic virus 35S (CaMV 35S) or rice actin (Act1) promoters were developed by particle gun bombardment of callus, suspension cells and cormel slices. Leaves of transgenic plants regenerated from cormel slices frequently exhibited a pattern of horizontal stripes following histochemical staining for β-glucuronidase (GUS) expression indicating the development of a chimeric transgenic plant. In comparison, leaves from transgenic plants regenerated from suspension cells were typically more uniform for GUS expression confirming that it was beneficial to use suspension cells rather than cormel slices for bombardment. In developing disease resistant plants it is critical that the transgene be expressed in a particular plant tissue and at a level necessary to confer resistance against the invading pathogen. Suspension cells were bombarded with gusA under various promoters to evaluate the effect of each promoter on levels of GUS expression. Transient GUS expression was evaluated 48 hr after bombardment and showed that levels of GUS expression were higher with dicot-associated promoters as compared to monocot-derived promoters. These results were confirmed by analyzing GUS expression in leaves of transgenic plants that had been stably transformed. Expression of the gusA gene under the CaMV 35S promoter in transgenic Gladiolus plants was used bo evaluate tissue-specific expression of GUS in various plant tissues. It was determined that the majority of the transgenic plants continued to express GUS in their leaves, roots and basal callus one year after they had been bombarded. Results from fluorescent 4-methyl umbelliferyl glucuronide (MUG) assays used to quantitate the specific activity of GUS confirmed the histochemical GUS expression in leaves. Many flowering bulb crops are multiplied by vegetative propagation rather than seed resulting in transfer of viruses to subsequent generations. Bean yellow mosaic virus (BYMV) is ubiquitous in Gladiolus. Transgenic Gladiolus plants containing either the BYMV coat protein or its antisense orientation under the enhanced CaMV 35S promoter were developed. These transgenic plants expressed various levels of the BYMV coat protein. Levels of expression were generally higher by plants grown in the greenhouse compared to in vitro. The BYMV coat protein was expressed after two seasons of dormancy. The technology described will be applied to the development of virus resistant floral bulb crops.
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