Molecular cloning of the complete 11S seed storage protein gene of Coffea arabica and promoter analysis in transgenic tobacco plants

In this paper, we present the complete nucleotide sequence of the csp1 gene from Coffea arabica coding for the 11S-globulin seed storage protein. To i nvestigate the sequences responsible for the regulated expression of this s eed-specific coffee storage protein gene, about 1 kb of the 5'-upstream reg ion from the csp1 gene was isolated using inverse polymerase chain reaction (IPCR) and then sequenced. Several DNA boxes were found in this coffee seq uence that had similarity to those previously identified as being essential for grain (endosperm) specific expression in other plants. To study the ab ility of this sequence to direct grain-specific expression, the whole fragm ent, as well as a series of 5' deletions, was fused to the reporter gene be ta-glucuronidase (uidA) and analysed in transgenic Nicotiann tabacum plants . GUS measurements showed that all the deletions of the csp1 promoter direc ted the expression of the reporter gene in tobacco grain but not in the oth er tissues examined. GUS activities also revealed that the csp1 promoter co nstructs function as very strong promoters by comparison to the strength of the cauliflower mosaic virus (CaMV) 35S promoter. Therefore, this 11S prom oter could represent a useful tool to change the expression of targeted gen es in the grain of transgenic coffee plants. (C) Elsevier, Paris.