Endosperm-specific activity of a storage protein gene promoter in transgenic wheat seed

The characterization of the promoter of a wheat (Triticum aestivum) cv, Cheyenne high molecular weight glutenin subunit (HMW subunit) gene, Glu-1D-1 is reported. The nucleotide sequence of the promoter from position -1191 to -650 with respect to the transcription start site was determined, to add to that already determined. Analysis of this region of the promoter revealed the presence of an additional copy of part of the primary enhancer sequence and sequences related to regulatory elements present in other wheat seed protein genes. A chimaeric gene was constructed comprising the 5' flanking region of the Glu-1D-1 gene from position -1191 to +58, the coding region of the UidA (Gus) gene, and the nopaline synthase (Nos) gene terminator. This chimaeric gene was introduced into wheat (Triticum durum cv. Ofanto) by particle bombardment of inflorescence explants. Two independent transgenic lines were produced, and both showed expression of the Gus gene specifically in the endosperm during mid-development (first detected 10-12 d after anthesis), Histochemical analysis of homozygous T-2 seed confirmed this pattern of expression, and showed that expression was initiated first in the central lobes of the starchy endosperm, and then spread throughout the endosperm tissue, while no expression was detected in the aleurone layer. Native HMW subunit protein was detectable by Western analysis 12-14 d after anthesis, consistent with concurrent onset of activity of the native and introduced HMW subunit gene promoters.