Co-ordinate expression of β- and δ-zeins in transgenic tobacco

Abstract

Two classes of alcohol soluble seed storage proteins found in the endosperm of maize contain unusually high levels of cysteine and methionine. These two proteins, the beta (β) and delta (δ) zeins, have been introduced into plants with the expectation of improving the sulfur nutritional content of various plants. Traditional methods of expressing multiple transgenes in plants include: (a) crossing transgenic plants which contain the genes of interest, (b) co-transforming the transgenes, and (c) successive retransformation. Co-ordinate expression of transgenes is not always successful with these traditional methods. We have co-ordinately expressed the β- and δ-zein proteins with the use of a synthetic self-hydrolyzing 2A peptide sequence utilized by a number of viruses [J. Gen. Virol. 82 (2001) 1013]. The β and δ zeins were fused with a 20 amino acid synthetic 2A peptide sequence between them. This β-zein–2A–δ-zein construct was introduced into tobacco. Western analysis indicates that tobacco plants containing this transgene accumulate both β- and δ-zein protein. The 2A peptide sequence cleaves correctly allowing the β- and δ-zein proteins to accumulate. Protein bodies are observed in these transgenic plants. This technology allows two genes to be expressed in one cassette, under the control of the same promoter, eliminating the traditional need for crossing or co-transformation.