Annual Progress Report on Malting Barley Research March, 2002

51
BARLEY TRANSFORMATION
G.J. Muehlbauer, C.A. Mackintosh, D.T. Tuong and N. Al-Saady
Department of Agronomy and Plant Genetics, University of Minnesota
St. Paul, MN 55108
Summary
1. We have modified a protocol for transforming the elite barley variety Conlon in our
laboratory. 1500, 1105 and 40 callus pieces have been bombarded with gold particles
coated with DNA encoding the following antifungal proteins (AFP): a wheat thaumatinlike
protein 1 (tlp-1), a barley chitinase and a barley glucanase. Many regenerating
plantlets are in the final stages of tissue culture. Further bombardments using the barley
glucanase and other AFP genes are on-going.
2. We have obtained affinity purified polyclonal antibodies to the following proteins; a
barley ribosome inactivating protein (RIP), a wheat tlp-1, a barley chitinase and a
barley glucanase. Protocols for using these antibodies in Western blotting to determine
AFP expression are being developed using transgenic wheat. Thus, we will be able to
determine the presence of these proteins in our transgenic barley as soon as the plants are
growing in soil.
3. We are developing in vitro fungal growth assays in order to identify antifungal
properties of transgenic barley seedlings. This will allow us to determine potential
antifungal effects before the plants reach flowering, the growth stage at which Fusarium
graminearum-glasshouse disease screening occurs.
4. We have continued to characterize barley plants transformed with the GUS gene
driven by the sugarcane badnavirus (ScBV) promoter. As described in the AMBA 2000-
2001 progress report, we developed twelve transgenic lines of barley expressing GUS
driven by the ScBV promoter. In the T1 generation we showed that the ScBV promoter
drives expression in all barley tissues in all twelve transgenic lines. Our work within the
last year was directed at determining the stability of GUS expression over generations.
Therefore, we have evaluated T2 generation lines. In the T2 generation, GUS was
expressed in eight of the 12 transgenic lines and appeared to be silenced in the other four
lines.
Introduction
Genetic transformation promises to be a powerful tool for improvement of crop plants.
Genetic manipulations that are not possible using conventional breeding methods are
attainable with plant transformation. Barley transformation has been accomplished via
particle bombardment into immature zygotic embryos, callus-derived immature embryos
and microspore-derived embryos (Wan and Lemaux, 1994).
Fusarium head blight (FHB), a fungal disease of small grain crops caused by Fusarium
graminearum, threatens to reduce barley to an economically unviable crop in Minnesota
and many other states. Currently, large breeding efforts at the University of Minnesota
and around the country are focused on developing FHB resistant cultivars. However,