A CHIMERIC GENE ENCODING THE METHIONINE-RICH 2S ALBUMIN OF THE BRAZILNUT (BERTHOLLETIA-EXCELSA HBK) IS STABLY EXPRESSED AND INHERITED IN TRANSGENIC GRAIN LEGUMES

The coding region of the 2S albumin gene of Brazil nut (Bertholletia e xcelsa H.B.K.) was completely synthesized, placed under control of the cauliflower mosaic virus (CaMV) 35S promoter and inserted into the bi nary vector plasmid pGSGLUC1, thus giving rise to pGSGLUC1-2S. This wa s used for transformation of tobacco (Nicotiana tabacum L. cv. Petit H avanna) and of the grain legume Vicia narbonensis L., mediated by the supervirulent Agrobacterium tumefaciens strain EHA 101. Putative trans formants were selected by screening for neomycin phosphotransferase (N PT II) and beta-glucuronidase (GUS) activities. Transgenic plants were grown until flowering and fruiting occurred. The presence of the fore ign gene was confirmed by Southern analysis. GUS activity was found in all organs of the regenerated transgenic tobacco and legume plants, i ncluding the seeds. In the legume, the highest expression levels of th e CaMV 35S promoter-controlled 2S albumin gene were observed in leaves and roots. 2S albumin was localized in the vacuoles of leaf mesophyll cells of transgenic tobacco. The Brazil nut protein was present in th e 2S fraction after gel filtration chromatography of the legume seed p roteins and could be clearly identified by immunoblotting. analysis of seeds from the R(2) progenies of the legume and of transgenic tobacco plants revealed Mendelian inheritance of the foreign gene. Agrobacter ium rhizogenes strain RifR 15834 harbouring the binary vector pGSGLUC1 -2S was also used to transform Pisum sativum L. and Vicia faba L. Hair y roots expressed the 2S albumin-specific gene. Several shoots were ra ised but they never completely rooted and no fertile plants were obtai ned from these transformants.