Isolation of two storage protein promoters from Hordeum chilense and characterization of their expression patterns in transgenic wheat
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Many plant genetic engineering applications require spatial expression of genes which in turn depends upon the availability of tissue and organ specific promoters. Wild plant species related to cultivated crops are an important source of genetic variability, not only for trait genes but also for regulatory sequences such as promoters. In the present work two new storage protein promoters cloned from the hordein type B and D genes of Hordeum chilense (Roem. et Schulz) are reported. The promoter of the B hordein gene contains the ‘prolamin box’, while the promoter of the D hordein shows the ‘High Molecular Weight (HMW) enhancer’ characteristic of the high molecular weight glutenin subunit (HMW-GS) promoters of wheat. The fragments of DNA corresponding to these promoters were fused upstream of the gusA gene in order to characterize their activities in transgenic bread wheat by histochemical staining and quantitative real time PCR (QRT-PCR) in different tissues and organs. Both promoters showed expression of the gusA gene in seeds but not in other tissues and organs. Both promoters also showed well-differentiated expression patterns in the endosperm. The promoter of the B hordein reached a maximum level of transcript abundance at 23 days post anthesis (DPA), while the promoter of the D hordein gave rise to high levels of transcript up to 28 DPA. With these two new promoters, the availability of sequences that direct endosperm-specific expression of genes of interest in the grain of wheat or other cereals is expanded.
KeywordsCereal transformation Hordeins Seed promoter Quantitative real time PCR
The authors acknowledge funding by the Spanish C.I.C.Y.T. (project AGL2004-03361-C02-2). We thank Dr. G. Dorado for his assistance with primer designing. Technical assistance by Ana García is also acknowledged.
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