Identification and characterization of a novel barley gene that is ABA-inducible and expressed specifically in embryo and aleurone

Abstract

barley cDNA library prepared from mRNAs of the ABA- Abstract A screening of a cDNA library of abscisic acid (ABA)-treated treated matured aleurone, which is rapidly induced by ABA. barley aleurone using a polyclonal anti-idiotypic antibody that A lgt22A phage library was constructed using mRNA isol- had been made against an anti-ABA monoclonal antibody ated from ABA-treated barley aleurone layers and a resulted in the isolation of a cDNA clone aba45. Northern blot Superscript lgt22A cDNA construction kit (GIBCO/BRL). analysis showed that aba45 was up-regulated by ABA and Aleurone layers were prepared from mature seeds of barley down-regulated by gibberellin. Aba45 mRNA was not detect- (Hordeum vulgare cv. Himalaya) and incubated with 100 mM able in barley roots, stems and leaves and was most abundant ABA for 48 h in the dark as described previously (Hill et al., in developing aleurone and embryo. Analysis of the 5ae genomic 1995). Mice were immunized four times intraperitoneally with sequence of aba45, isolated using a nested PCR procedure, 100 mg anti-ABA monoclonal antibody (IDETEK, Inc.) in revealed a conserved ABA response complex that consists phosphate buVer saline. The crude serum was withdrawn and the total IgG antibody (AB2) was purified using Protein A