Location: Cereal Crops Research
2023 Annual Report
Objectives
Objective 1. Resolve the genetic basis of dormancy and Preharvest Sprouting (PHS) in malting barley.
Objective 1.1: Demonstrate that domestication of barley for improved malting has increased the frequency of mutations within genes that affect dormancy and PHS.
Objective 2. Evaluate and report intrinsic malting quality parameters of commercially viable barley cultivars as part of a Congressionally-directed mission of service (non-hypothesis driven).
Objective 2.1: Evaluate and report intrinsic barley malting quality parameters used to guide breeder selection for superior variety development and evaluation of new varieties.
Approach
We plan to test preharvest sprouting on three unique panels of barley that include lines of contemporary, heritage, and globally diverse origin to reveal the extent of the genetic contribution to dormancy (or nondormancy) together with the screening of these lines for precocious germination on intact heads with the goal of developing genetic resources to reduce preharvest sprouting in North American varieties. The first phase of the approach will examine associations between PHS susceptible phenotypes and allelic variability by using each of the three panels as subjects of genotypic and phenotypic evaluation to 1) to assess sequence variation through a targeted sequencing approach of genes associated with PHS and 2) to develop preharvest sprout scores using mist treatments and germination assays. The second phase will employ a gene discovery-based approach using GWAS on genotyped lines in each of the three populations to identify SNPs (or combinations of SNPs) that can uniquely identify plants carrying alleles that influence PHS related traits. These QTL will fortify our developing panel of novel alleles with additional mutations associated with PHS. Finally, molecular markers targeting SNPs tightly associated with PHS will be developed with the intent of deployment for Marker Assisted Selection (MAS) in malting barley breeding programs.
Processing and test procedures are based on the ASBC Methods of Analysis, with some slight modifications and improvements. A base calculation of steep time is generated by determining the average kernel weight (mg) of the sample and using an empirically-determined relationship to yield a base steep time in hours. Slight final adjustments to moisture levels are made prior to moving the samples to the germinator. At this point, we treat all of the submissions equally, using a standardized malting protocol that has been used reliably since 1998. Samples are germinated at 16ºC, 100% humidity, with intermittent turning. An intermediate weight for each sample is taken and a final water adjustment to maintain 45% moisture is made if necessary. After 5 days of germination, the “green malt” is moved to the kiln and the samples are dried for 24 hours, starting with a temperature of 49ºC for 10 h, and rising to 85ºC for last 3 hours. Once the samples are steeped, malted, kilned, and cleaned (rootlets and emergent acrospires removed), they are stored to allow moisture equilibration and sample aging. Samples are subjected to industry standard protocols by the American Society of Brewing Chemists (ASBC). Malts are ground and mashed (ASBC Malt-4) and analyzed for % extract (Malt-4), soluble protein (Wort-17), color and clarity (Wort-9), free amino nitrogen content (Wort-12) and beta-glucan levels (Wort-18). Malt grist is extracted into salt water at 20ºC and the diastatic power (Malt-6) and alpha-amylase activities (Malt-7) are determined. Total nitrogen contents of the barley are determined using FOSS Nova NIT and malt are determined on a LECO Corp. FP528 Nitrogen Analyzer utilizing the Dumas total combustion method (Malt-8).
Progress Report
Progress was made on both objectives. Objective 1: Previous work has highlighted the effect of two genes that control germination: one gene allows barley to resist preharvest sprouting (PHS) by increasing a seed’s tendency to remain dormant, while the other gene has an opposite effect and causes susceptibility to preharvest sprouting by driving precocious and rampant germination prior to harvest. Neither precocious germination nor pronounced seed dormancy are desirable traits in malting barley as they both are negatively correlated with the quality of malt made with that barley. However, current research, by ARS scientists in Madison, Wisconsin, using high density genotyping techniques and DNA sequencing, has discovered that a specific combination of genes allows for production of high-quality malt targeted to the craft malting/beer industry from barley that both germinates well and is resistant to PHS. Scientists in Madison micromalted and analyzed the malt characteristics from all the barley lines that were used in the gene sequencing survey and confirmed that these genes driving aggressive germination in barley are also strongly correlated to malt quality characteristics.
ARS researchers in Madison, Wisconsin, are expanding their scope towards the understanding and prevention of grain PHS and also overall grain quality through the use of spectral analysis to predict characteristics of barley grains. Toward that goal, ARS researchers in Madison, Wisconsin, have incorporated both lab-based spectral sensors and Unmanned Aerial Vehicles (UAV) equipped with spectral sensors for both monitoring the quality of seed as it develops in the production fields, as well as predicting the potential of the grain harvested from these fields to produce high quality malt in malt houses. For example, lab-based work with hyperspectral sensors demonstrated the potential to predict whether a seed has begun germinating or retains a dormant state and can do so at least 4 days before any visual signs of the trait is observed. Furthermore, the same type of analysis using the hyperspectral sensors can also roughly predict the level of an enzyme produced during malting that is of critical importance to the brewing industry, namely alpha amylase. To complement this approach, ARS researchers in Madison, Wisconsin, have installed a PHS nursery at a research station where barley varieties will be induced to preharvest sprout using irrigation, evaluated for levels of PHS resistance, and used a subject for drone inspection to assess whether preharvest sprouted grain can be determined remotely while the grain is still in the field. ARS researchers in Madison, Wisconsin, will further develop the use of spectral imaging for both drone-monitoring of barley research plots and ground-based monitoring of malting with the goal of using the data collected to predict malt quality based on growth and spectral characteristics from the maturing grain.
Determining the overall quality of malt from barley is expensive and requires trained scientists operating technical lab equipment to produce. The labor and costs of this analysis would be greatly reduced if the quality of malt could be determined using spectral imaging. ARS scientists in Madison, Wisconsin, have collaborated with researchers at the University of Wisconsin to evaluate different computational methods to analyze the spectral data to determine which approach provides most accurate method to predict malt quality.
ARS researchers at Madison, Wisconsin, have previously reported on utilizing wild barley species as a resource to improve the nutrition of cultivated barley, and uncovered genetic markers controlling beta glucan content. Building upon the work in wild barley, an ARS researchers in Madison, Wisconsin, has identified and reported on genetic markers that affect the overall grain content of key vitamins and minerals such as zinc, calcium, manganese and iron; the deficiencies of which (zinc and iron) contribute to hidden hunger in developing countries.
In support of Objective 2: ARS researchers in Madison, Wisconsin, worked diligently to progress through the highest priority barley malt quality samples to meet stakeholder deadlines. The lab continues to progress its agenda of overall capacity expansion and increased throughput, primarily through the addition of instrumentation and subsequent fortification of lab personnel. To date, the lab has processed nearly all samples submitted from the public sector breeder’s 2022 Crop Year (CY) material: It has received 4385 barleys and all have been cleaned and malted. Quality analyses have been completed on 4187 malts. In collaboration with a stakeholder’s pilot barley program, the lab has completed 80 of their top priority samples for their group. A table with cumulative results for the 2022 CY will be posted on a website after completion of analyses.
Accomplishments
1. Novel method for identifying barley lines resistant to pre-harvest sprouting. ARS researchers in Madison, Wisconson, identified a specific combination of genes by DNA sequencing allowing for identification of barley lines resistant to Pre-Harvest Sprouting. Production of high-quality malt targeted to the craft malting/beer industry from barley that germinates well as well as being resistant to PHS is now possible using these markers.
Review Publications
Abendroth, J.A., Sallam, A.H., Steffenson, B.J., Vinje, M.A., Mahalingam, R., Walling, J.G. 2022. Identification of genomic loci controlling grain macro and micronutrients variation in a Wild Barley (H. vulgare spp spontaneum) Diversity Panel. Agronomy Journal. 12(11). Article 2839. https://doi.org/10.3390/agronomy12112839.
Rooney, T.E., Sweeney, D.W., Kunze, K.H., Sorrells, M.E., Walling, J.G. 2023. Malting quality and preharvest sprouting traits are genetically correlated in spring malting barley. Theoretical and Applied Genetics. 136. Article 59. https://doi.org/10.1007/s00122-023-04257-6.
