Fig. 1.

Experimental design and gene constructs. (a) The selection and propagation process of the plants and seeds used in this study. (b) The binary vectors used for Agrobacterium-mediated transformation are shown. The regulatory elements include: 7S-P (7S soybean β-conglycinin promoter), TEV (tobacco etch virus translational enhancer element), hTG (human thyroglobulin gene), hTG-SP (hTG signal peptide), 35S-T (35S cauliflower mosaic virus terminator element), Gly-SP (soybean glycinin signal peptide), hMBP-Sigma (human myelin basic protein fused to Reovirus Sigma 1 protein), 11S-P (soybean 11S glycinin promoter), mSEB (mutant nontoxic staphylococcal enterotoxin B gene), NOS-P (nopaline synthase promoter), BAR (phosphinothricin acetyltransferase gene) and NOS-T (nopaline synthase terminator element). Arrows indicate orientation of cassettes relative to the right border (RB) and left border (LB) sequences. Regulatory elements and genes are not drawn to scale. Molecular characterization of transgenic events. (c) Duplex PCR of the nine progeny seeds from the indicated transformation events. wt: nontransgenic (negative control); +: plasmid DNA (positive control). Arrows indicate amplified DNA fragments derived from the specific gene of interest as well as vegetative storage protein gene (VSP) following separation in agarose gels. Sizes of molecular weight markers are shown in base pairs. (d) Western blots of total seed protein derived from the transgenic progenies shown in (c). Arrows indicate the hTG, mSEB and hMBP-Sigma immunoreactive proteins. Sizes of molecular weight standards are shown as kDa. Positive controls (+) are purified hTG (Cal Biochem), E. coli-derived mSEB, and soy-derived hMBP-Sigma from a higher expressing line