Abstract
Conditions have been optimized for the routine isolation and transfection of protoplasts from developing endosperm, aleurone, leaves and roots from barley ( Hordeum vulgare L.) and wheat ( Triticum aestivum L.). Transient expression of the β-glucuronidase (GUS) reporter gene under the control of the cauliflower mosaic virus 35S promoter was used to evaluate the best procedure for each tissue. Electroporation was ∼300% more efficient than polyethyleneglycol (PEG) mediating 35S-GUS transient expression in leaf and root protoplasts while endosperm protoplasts had to be PEG transfected since even the mildest electroporation parameters tested (250 V/cm, 200 μF, 10 ms) were lethal for them. Protoplasts isolated from dry aleurone showed the highest viability (approximately 90%), but GUS activity after PEG transfection was 50% of that obtained with endosperm protoplasts. Conditions optimized for a given tissue of a barley cultivar such as Bomi, were readily applied to other barley cultivars and to other cereal species.
Published Version
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