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The use of dwarfing genes has resulted in the most significant improvements in yield and adaptation in cereal crops. The allelic dwarfing gene sdw1/denso has been used throughout the world to develop commercial barley varieties. The sdw1 gene has never been used successfully for malting barley, but only for a large number of feed varieties. One of the gibberellin 20-oxidase genes (Hv20ox₂) was identified as the candidate gene for sdw1/denso. Semi-quantitative real-time RT-PCR revealed that Hv20ox₂ was expressed at different levels in various organs of barley. Transcriptional levels were reduced in leaf blade, sheath, stem and rachis tissue in the barley variety Baudin with the denso gene. Subsequently, the relative expression levels of Hv20ox₂ were determined by quantitative real-time RT-PCR in a doubled haploid population and mapped as a quantitative trait. A single expression quantitative trait locus (eQTL) was identified and mapped to its structural gene region on chromosome 3H. The eQTL was co-located with QTLs for yield, height, development score, hectolitre weight and grain plumpness. The expression level of Hv20ox₂ was reduced fourfold in the denso mutant, but around 60-fold in the sdw1 mutant, compared to the control variety. The reduced expression level of Hv20ox₂ enhanced grain yield by increasing the number of effective tillers, but had negative effects on grain and malting quality. The sdw1 gene can be used only in feed barley due to its severe reduction of Hv20ox₂ expression. The gene expression marker for Hv20ox₂ can be used to distinguish different alleles of sdw1/denso.