BETA-GLUCURONIDASE GENE-EXPRESSION AND MESSENGER-RNA STABILITY IN OATPROTOPLASTS

Protoplasts derived from oat (Avena sativa L.) suspension culture cell s (7 days after subculturing) were electroporated with plasmid DNA con taining the Escherichia coli uidA gene encoding the beta-glucuronidase reporter enzyme. Consistently high enzyme activity was observed with electroporation conditions of 500 muF and 1125 volts/cm. Enzyme activi ty and mRNA accumulation time courses were determined. The maximum enz yme activity was detected at 24 hours after electroporation, while the maximum mRNA level was detected at 12 hours after electroporation. Be ta-glucuronidase mRNA was in vitro synthesized with and without a 5' m ethylated cap and then electroporated into protoplasts. Only capped mR NA produced significant enzyme activity. By electroporating radiolabel ed, in vitro synthesized mRNA, the beta-glucuronidase mRNA half-life w as estimated to be approximately 35 minutes in oat protoplasts.