Higher-level accumulation of foreign gene products in transgenic rice seeds by the callus-specific selection system

Transgenic rice plants that accumulate the hypocholesterolemic pentapeptide lactostatin (IIAEK) as a fusion protein with the endosperm seed storage protein glutelin were developed. Plants were selected from Agrobacterium -mediated transformants using a mutated-rice acetolactate synthase ( mALS ) marker protein expressed under the control of the rice callus-specific promoter (CSP) (CSP: mALS ). Lactostatin accumulation levels were compared in mature seeds of 78 independent transgenic rice lines selected for either the CSP: mALS gene cassette or hygromycin phosphotransferase (HPT) under the control of the CaMV 35S promoter (35S: HPT ). Transgenic rice seeds harboring a CSP: mALS gene cassette accumulated more (approximately 120–200% on average) lactostatin than those with a 35S: HPT gene cassette, and accumulation was largely independent of transgene copy number. Furthermore, transgenic rice seeds that were selected by HPT under the control of CSP (CSP: HPT ) also accumulated more lactostatin (approximately 160% on average) than transgenic rice seeds with the 35S: HPT gene cassette. These results indicate that high-value bioactive peptides such as lactostatin can be produced at higher levels using callus-specific selection than by conventional constitutive selection. Specific expression of a marker gene at the selection stage may thus result in increased target peptide accumulation levels in seeds.