Copyright (c) 2009 All rights reserved. http://works.bepress.com/agnelo_furtado Recent documents in Agnelo Furtado en-us Wed, 28 Oct 2009 23:19:52 PDT 3600 http://works.bepress.com/agnelo_furtado/49 http://works.bepress.com/agnelo_furtado/49 Tue, 27 Oct 2009 22:51:58 PDT Agnelo Furtado http://works.bepress.com/agnelo_furtado/48 http://works.bepress.com/agnelo_furtado/48 Tue, 20 Oct 2009 22:50:33 PDT Robert J. Henry http://works.bepress.com/agnelo_furtado/47 http://works.bepress.com/agnelo_furtado/47 Tue, 01 Sep 2009 23:20:20 PDT The bi-functional alpha amylase subtilisin inhibitor (ASI), an endogenous inhibitor synthesised in the barley grain, inhibits high pI-group of alpha-amylases and the bacterial protein subtilisin. The expression of the asi gene in developing barley grain is thought to be under control of the hormones abscisic acid (ABA) and giberellic acid (GA), a control which is antagonistic to that of alpha-amylase. Such controlled expression of ASI and alpha-amylase ensures that starch is either accumulated during grain filling or is digested during seed germination. The levels of ASI as compared to total seed proteins remain relatively stable in response to various climatic conditions. This suggests that the promoter and possibly the 3`-untranslated region of the asi could be used to control stable expression of trans-genes in the barley grain and possibly other cereals. The promoter and the 3'-untranslated region of asi have been isolated and a number of putative DNA-elements, which are known to regulate expression of other genes in endosperm tissue, have been identified. Functional analysis of progressive 5'-deletions of the asi-promoter fused to the green fluorescent protein gene (gfp) will be studied to identify regions controlling hormonal regulation and endosperm specific expression. The 3'-untranslated region of asi will also be studied to determine it's role in hormonal control or message stability. Agnelo Furtado http://works.bepress.com/agnelo_furtado/46 http://works.bepress.com/agnelo_furtado/46 Tue, 11 Aug 2009 23:31:14 PDT The bifunctional alpha-amylase subtilisin inhibitor (BASI) from barley is a bifunctional protein with roles in inhibition of proteases and of endogenous alpha-amylase. The subtilisin inhibitor activity has been attributed to a defence role for this protein in barley. The specific inhibition of barley alpha-amylases by this protein has been previously explained as a role associated with the control of starch mobilisation in germinating cereals. It has long been assumed that this protein was expressed in aleurone and endosperm of barley. We recently isolated the promoter of the asi gene and demonstrated pericarp specific expression of GFP with this promoter. Expression of this gene specifically in the pericarp of developing barley suggests that the protein is primarily associated with defence of the seed against pathogens. The action of the protein in specifically inhibiting barley alpha-amylase may be associated with denial of fungal access to seed starch reserves. Robert J. Henry http://works.bepress.com/agnelo_furtado/45 http://works.bepress.com/agnelo_furtado/45 Tue, 11 Aug 2009 23:31:13 PDT Robert J. Henry http://works.bepress.com/agnelo_furtado/44 http://works.bepress.com/agnelo_furtado/44 Tue, 11 Aug 2009 23:31:12 PDT Agnelo Furtado http://works.bepress.com/agnelo_furtado/43 http://works.bepress.com/agnelo_furtado/43 Tue, 11 Aug 2009 23:31:12 PDT Robert J. Henry http://works.bepress.com/agnelo_furtado/42 http://works.bepress.com/agnelo_furtado/42 Tue, 11 Aug 2009 23:31:10 PDT Robert J. Henry http://works.bepress.com/agnelo_furtado/41 http://works.bepress.com/agnelo_furtado/41 Mon, 03 Aug 2009 14:41:00 PDT The bifunctional α-amylase/subtilisin inhibitor (BASI) is an abundant protein in barley seeds, proposed to play multiple and apparently diverse roles in regulation of starch hydrolysis and in seed defence against pathogens. In the Triticeae, the protein has evolved the ability to specifically inhibit the main group of α-amylases expressed during germination of barley and encoded by the amy1 gene family found only in the Triticeae. The expression of the asi gene that encodes BASI has been reported to be controlled by the hormones abscisic acid (ABA) and gibberellic acid (GA). Despite many studies at the gene and protein level, the function of this gene in the plant remains unclear. In this study, the 5'-flanking region (1033 bp, 1033-asi promoter) and the 3'-flanking region (655 bp) of the asi gene were isolated and characterised. The 1033-asi promoter sequence showed homology to a number of ciselements that play a role in ABA and GA regulated expression of other genes. With a green fluorescent protein gene (gfp) as reporter, the 1033-asi promoter was studied for spatial, temporal and hormonal control of gene expression. The 1033-asi promoter and its deletions direct transient gfp expression in the pericarp and at low levels in mature aleurone cells, and this expression is not regulated by ABA or GA. In transgenic barley plants, the 1033-asi promoter directed tissue-specific expression of the gfp gene in developing grain and germinating grain but not in roots or leaves. In developing grain, expression of gfp was observed specifically in the pericarp, the vascular tissue, the nucellar projection cells and the endosperm transfer cells and the hormones ABA or GA did not regulate this expression. In mature germinating grain gfp expression was observed in the embryo but not in aleurone or starchy endosperm. However, GA induced gfp expression in the aleurone of mature imbibed seeds from which the embryo had been removed. Expression in maternal rather than endosperm tissues of the grain suggests that earlier widespread assumptions that the protein is expressed largely in the endosperm may have been largely based on analysis of mixed grain tissues. This novel pattern of expression suggests that both activities of the protein may be primarily involved in seed defence in the peripheral tissues of the seed. Agnelo Furtado http://works.bepress.com/agnelo_furtado/40 http://works.bepress.com/agnelo_furtado/40 Mon, 03 Aug 2009 14:40:59 PDT The present invention provides a novel promoter that is functional in the seeds of plants, in particular the endosperm and/or aleurone and/or the scutellum. Preferably, the isolated promoter sequence of the invention further modulates the expression of a structural gene in response to the phytohormones ABA and/or GA. Alternatively or in addition, the promoter sequence of the present invention is capable of being repressed or otherwise down-regulated in response to GA. The present invention further encompasses genetic constructs capable of expressing a structural gene operably under the control of the inventive promoter sequence, and transgenic plants carrying the genetic constructs. The promoter sequence of the present invention is particularly useful for modifying the seed traits of plants. Agnelo Furtado