Location: Cereal Crops Research2012 Annual Report
1a. Objectives (from AD-416):
Objective 1: To assess allelic variation in Agl1, the gene encoding a-glucosidase, in barley (Hordeum vulgare ssp. vulgare) germplasm adapted to North America and in selected Hordeum vulgare ssp. spontaneaum accessions currently being used to introgress disease resistance genes into North American barley germplasm. Objective 2: To determine the potential contributions of isoamylase to the production of fermentable sugars during mashing and, if isoamylase contributes to fermentable sugar production, assess allelic variation in selected barley populations. Objective 3: To determine metabolic profiles of worts or 20ºC water extracts to determine which metabolites distinguish the best malting barleys from those of lesser malting quality.
1b. Approach (from AD-416):
Metabolic profiling of malts will be conducted using standard methods of derivatization, gas chromatography and mass spectral detection/ identification of sample components followed by multivariate analysis of data to identify metabolites that best define the trait "malting quality". Metabolite identification will be done using several mass spectra databases including the 7th edition of Wiley, Essential Oils/Flavor and Fragrance and my lab's personal database. Examination of enzyme contributions to fermentable sugar production will be by use of an immunological probe and by characterizing the recombinant enzyme in a heterologous expression system. Assessment of allelic variation of carbohydrases will be via PCR amplification of expressed mRNA followed by sequence analysis and exploration of functional consequences of documented polymorphisms.
3. Progress Report:
Analysis of expression of a carbohydrase encoding gene included determination of tissues the promoter targeted, development of deletion cassettes to determine regions of promoter necessary for expression, and initiating analysis of promoter regions. Promoter analysis and timing of expression of the gene and its product showed similarities to major storage proteins of barley; hence, work was conducted to directly compare mobilization of the protein during germination and malting to that of seed storage proteins. We completed analysis of the effect of allelic variation in a key carbohydrase on the ability of malts to produce fermentable sugars during mashing. Metabolic profiling of oats recovering from low temperature stress was completed. Metabolic profiling of two barley populations resulting from crosses of winter barleys with enhanced low temperature tolerance with high malting quality barleys was initiated. Two environments were planted, sampled twice in the fall/winter, one environment was sampled during spring regrowth, and spring survival counts were conducted. Sample analysis was initiated and approximately one-third are completed. Mature seeds will be harvested and analyzed for malt quality resulting in identification of metabolite qualitative trait loci (QTL) coincident with malt quality QTL. The carbohydrase genes studied expanded from Agl1 to include bmy1, which is that reported on here, as work on agl1 was completed previously.
1. Evaluation of the utility of a molecular marker for enhanced malt quality. The international development of a molecular marker for use in efficient selection of high quality malting barleys has generated conflicting data on its general applicability to a broad spectrum of germplasm. ARS researchers at Madison, Wisconsin along with University of Wisconsin colleagues demonstrated that this specific marker is not useful in the selection of germplasm with enhanced ability to produce sugars during the mashing of malting barley germplasm with North American parentage. This is consistent with all other tests of this marker’s ability to predict malts of North American lineage with enhanced starch conversion regardless of the specific performance metric studied. The impact is that North American barley breeders will not expend limited resources on the use of this molecular marker as a general tool, but will limit its application to discrete introductions of new alleles from international germplasm. This allows limited resources to be better deployed on markers with proven utility in their germplasm collections.
Duke, S.H., Henson, C.A. 2011. Tracking the progress of wort sugar production during congress mashing with North American barley cultivars and comparisons to wort osmolyte concentrations and malt extract. Journal of American Society of Brewing Chemists. 69(4):200-213.