Targeted mutation of barley (1,3;1,4)-β-glucan synthases reveals complex relationships between the storage and cell wall polysaccharide content

Plant J. 2020 Nov;104(4):1009-1022. doi: 10.1111/tpj.14977. Epub 2020 Sep 29.


Barley (Hordeum vulgare L) grain is comparatively rich in (1,3;1,4)-β-glucan, a source of fermentable dietary fibre that protects against various human health conditions. However, low grain (1,3;1,4)-β-glucan content is preferred for brewing and distilling. We took a reverse genetics approach, using CRISPR/Cas9 to generate mutations in members of the Cellulose synthase-like (Csl) gene superfamily that encode known (HvCslF6 and HvCslH1) and putative (HvCslF3 and HvCslF9) (1,3;1,4)-β-glucan synthases. Resultant mutations ranged from single amino acid (aa) substitutions to frameshift mutations causing premature stop codons, and led to specific differences in grain morphology, composition and (1,3;1,4)-β-glucan content. (1,3;1,4)-β-Glucan was absent in the grain of cslf6 knockout lines, whereas cslf9 knockout lines had similar (1,3;1,4)-β-glucan content to wild-type (WT). However, cslf9 mutants showed changes in the abundance of other cell-wall-related monosaccharides compared with WT. Thousand grain weight (TGW), grain length, width and surface area were altered in cslf6 knockouts, and to a lesser extent TGW in cslf9 knockouts. cslf3 and cslh1 mutants had no effect on grain (1,3;1,4)-β-glucan content. Our data indicate that multiple members of the CslF/H family fulfil important functions during grain development but, with the exception of HvCslF6, do not impact the abundance of (1,3;1,4)-β-glucan in mature grain.

Keywords: (1,3;1,4)-β-glucan; CRISPR/Cas9; barley; cell walls; gene editing.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Wall / metabolism
  • Edible Grain
  • Gene Editing
  • Glucosyltransferases / genetics
  • Glucosyltransferases / metabolism
  • Hordeum / enzymology*
  • Hordeum / genetics
  • Mutagenesis, Site-Directed
  • Mutation
  • Plant Proteins / genetics
  • Plant Proteins / metabolism*
  • Plants, Genetically Modified
  • Polysaccharides / metabolism
  • beta-Glucans / metabolism*


  • Plant Proteins
  • Polysaccharides
  • beta-Glucans
  • beta-glucan, (1-3)(1-4)-
  • Glucosyltransferases
  • cellulose synthase
  • glucan synthase