Biolistic transformation, expression, and inheritance of bovine β-casein in soybean (Glycine max)

A 630-bp DNA fragment encoding a bovine milk protein, β-casein, was cloned into a seed-specific lectin promoter expression cassette and introduced into soybean somatic embryos via particle bombardment. Four hygromycin-resistant embryogenic cell lines were selected. All four lines were positive for the β-casein gene via Southern blot analysis; however, only one line did not show significant rearrangement of the β-casein gene and/or lectin promoter. The presence of the β-casein gene in the genome of the four plants regenerated from this culture was confirmed via Southern blot hybridization analysis. DNA blot experiments and progeny inheritance analysis indicated that the plants contain four to eight copies of the β-casein gene and that the insertion occurred at a single genetic locus. Bovine β-casein mRNA was highly expressed in developing cotyledons. A very low level of β-casein mRNA was found in leaf tissues although endogenous lectin transcripts were not found in the same samples. Bovine β-casein protein was expressed in cotyledons of transformed plants but was not detectable in leaf tissues of transformed plants by immunoblotting. Bovine β-casein produced in the individual developing seed of the first generation of transgenic plants migrated as two closely spaced bands very similar to those of pure bovine β-casein. This experiment represents the first report of the expression of a milk protein in soybean and opens the way for the general improvement of protein quality in soybean directed by seed-specific promoters.