Genotypic and phenotypic variation in beta-amylases from cultivated and wild barleys

Barley (Hordeum vulgare L.) beta-amylase is encoded by two genes; an endosperm-specific gene (Bmy1) and a ubiquitous gene (Bmy2). Markers based on polymorphisms in Bmy1 intron III have been proposed as candidates for use in marker assisted selection for beta-amylase activity and thermostability. Bmy1 intron III in 40 barley genotypes was sequenced and four alleles were identified based on four large insertion/deletions (indels). Due to the wide range of beta-amylase activity and thermostability observed among genotypes with the same Bmy1 intron III allele; these alleles are poor candidates for use as markers in marker assisted selection. RNA expression of Bmy1 and Bmy2 in developing grains at 17, 19, and 21 days after anthesis (DAA) was examined in two cultivated barleys (Legacy, Harrington) and two wild barleys (Ashqelon, PI 296978). Each genotype carries a different Bmy1 intron III allele. Bmy1 RNA expression in Ashqelon and PI 296897 had 2.5- to 3-fold higher Bmy1 RNA expression than Legacy or Harrington at 17, 19, and 21 DAA. Indels in Bmy1 intron III or in the Bmy1 promoter do not appear to affect Bmy1 RNA expression. The 503 bp upstream of Bmy1 is highly conserved among wild and cultivated barleys and may possibly contain the requisite transcription factor binding sites barleys and may possibly contain the requisite transcription factor binding sites necessary for Bmy1 transcription. The expression pattern of Bmy1 RNA indicates that Bmy1 may be under the control of seed storage protein transcription factors. PI 296897 had significantly higher Bmy2 RNA expression than the other three genotypes at 17, 19, and 21 DAA. However, the ratio of Bmy1 to Bmy2 RNA expression was 20,000 to over 100,000 for Legacy, Harrington, and Ashqelon at 17, 19, and 21 DAA and PI 296897 at 19 and 21 DAA. PI 296897 had 5,000 times more Bmy1 mRNA than Bmy2 mRNA in grains at 17 DAA. An attempt to identify the Bmy2 protein in grains from PI 296897 at 17 DAA and maturity was inconclusive. If the Bmy2 protein is present it is present in very small amounts. Therefore, the vast majority of beta-amylase activity observed in developing and mature grains can be attributed to the Bmy1 gene.