Submitted to: Crop Science
Publication Type: Peer reviewed journal
Publication Acceptance Date: 11/21/2001
Publication Date: 12/1/2002
Citation: DAHLEEN, L.S., BREGITZER, P.P. AN IMPROVED MEDIA SYSTEM FOR HIGH REGENERATION RATES FROM BARLEY IMMATURE EMBRYO-DERIVED CALLUS CULTURES OF COMMERCIAL CULTIVARS. CROP SCIENCE. 2002. VOL. 42:934-938. Interpretive Summary: Several important genetic techniques, such as insertion of new genes, require growth of plant cells on nutrient media and inducing them to regenerate many new plants. These techniques have been limited to a few unadapted barley varieties because most barley types do not regenerate enough plants on standard nutrient media. We have improved the nutrient composition so these techniques can be used on elite barley varieties that are adapted to North America. Ten varieties representing the types of barley grown in the US were compared for plant regeneration on standard and improved media. Increasing boric acid 7.5-fold and cutting the amount of iron in half, in addition to several other changes previously identified, increased the number of plants regenerated from all barley varieties. Average plant regeneration on the old standard media was two plants per petri dish, compared to an average of 29 plants regenerated on the improved media. These improvements mean that elite adapted cultivars can now be used for these genetic techniques, speeding the development of improved barley varieties.
Technical Abstract: Improving green plant regeneration from immature embryo-derived calli of modern commercial barley cultivars will facilitate their genetic transformation. Significant improvements in green plant regeneration were achieved with an improved media micronutrient composition. These improvements were identified based on the plant regeneration responses of 'Morex' and 'Harrington', the malting quality standard cultivars in North America, which regenerate few green plants when cultured on Murashige and Skoog- (MS) based media. Green plant regeneration response to various concentrations of eight micronutrients indicated that standard (MS) concentrations were appropriate for KH2PO4, NaMoO4, MnSO4, KI, CoCl2, and ZnSO4. Increased H3BO3 (0.75 mM) and decreased FeSO4 (0.05 mM) improved green plant regeneration. Changes in H3BO3 and FeSO4 concentrations were incorporated with other improvements reported previously, and the combined effect of all changes assessed by comparison to the regeneration responses of callus cultured on our previously-used MS-based media regime. We also assessed the effects of improvements on regeneration from callus cultures of seven elite breeding lines and cultivars, and of 'Golden Promise', which is relatively easy to transform in part because of its high rate of green plant regeneration. Green plant regeneration was improved for all cultivars and breeding lines, and increased from an average of two green plants per petri dish to 29 green plants per petri dish on improved media. Regeneration rates from all elite germplasm lines and cultivars on the improved media exceeded that of Golden Promise on the original media, indicating that poor plant regeneration no longer should be a limiting factor in the transformation of modern North American cultivars.