Transient expression of foreign genes introduced into barley endosperm protoplasts by PEG-mediated transfer or into intact endosperm tissue by microprojectile bombardment

Starchy endosperm protoplasts from developing barley (Hordeum vulgare) grains were isolated 8–15 days post-anthesis. These endosperm protoplasts were incubated in a hormone free medium containing sucrose as a carbon source and glutamine as a nitrogen source. In this medium the endosperm protoplasts remained viable for several days and starch synthesis was also observed. Transient expression of chloramphenicol acetyl transferase (CAT) and β-glucuronidase (GUS) reporter genes linked to the CaMV 35S promoter was detected 18–20 h after PEG induced DNA uptake. GUS fusions to two endosperm specific promoters, a wheat high molecular weight (HMW) glutenin and a barley chymotrypsin-inhibitor 2 were also functional in the endosperm protoplasts. Similarly, GUS activity could be detected when either the CaMV 35S or HMW glutenin promoters linked to the GUS gene were introduced into intact endosperm tissues of 15–20 days post-anthesis barley grains by microprojectile bombardment.