Optimisation of Tissue Culture Conditions for Transformation Studies Using Immature Embryos of Australian Barley Cultivars

Abstract

Eight Australian barley cultivars were tested for efficiency of embryonic callus initiation and plant regeneration, from immature embryo explants in tissue culture. Optimisation of tissue culture conditions was performed for cultivars Bandulla, Clipper, Schooner and Tallon in an attempt to increase regeneration frequencies to levels suitable for genetic engineering of barley. Variables tested were 2,4-D concentration, salt composition, carbon source and immature embryo explant. Optimal culture medium composition varied between cultivars. Shoot regeneration rates from culture of isolated scutellar tissues were low for all four cultivars. Halved, immature embryos produced most shoots for cultivars Clipper, Schooner and Tallon, whereas Bandulla performed best with entire immature embryo explants. Clipper (a malting barley) and Bandulla (a feed barley) are suggested as model Australian cultivars for transformation studies. Immature embryos of Bandulla produced an average of 5.3 shoots and Clipper 10.1 shoots per embryo under optimal conditions. Our results show that rates of somatic embryo and plant regeneration sufficient for use in transformation studies can be achieved for diverse Australian Barley cultivars, through systematic testing of a range of key variables including explant type and medium composition.