ISOLATION AND TRANSFORMATION OF RICE ALEURONE PROTOPLASTS

Protoplasts isolated,from the aleurone have been used extensively in m olecular studies focusing on hormone-mediated regulation of gene expre ssion in barley seed. To extend the use of aleurone protoplasts to oth er species, we have determined the conditions necessary for the isolat ion of protoplasts from rice aleurone layers of germinated seed. Many of the common cell wall degrading enzymes used in making protoplasts w ere tested for their ability to release protoplasts from rice aleurone layers. Cellulysin was found to be the most effective. Transformation of these aleurone protoplasts was accomplished using polyethylene gly col and DNA constructs containing the firefly luciferase reporter gene under the control of two different promoters were tested. Luciferase expression was 24-fold greater when the reporter gene was under the co ntrol of the CaMV 35S promoter than when the promoter from the alcohol dehydrogenase 1 gene was used. With the isolation and transformation of aleurone protoplasts from rice, it is now possible to investigate m olecular events occurring in this tissue during germination.