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

By Guillermo Garcia-Gimenez, Abdellah Barakate, Pauline Smith, Jennifer Stephens, Shi F. Khor, Monika Susanne Doblin, Pengfei Hao, Antony Bacic, Geoffrey B. Fincher, Rachel A. Burton, Robbie Waugh, Matthew R. Tucker, Kelly Houston

Posted 14 May 2020
bioRxiv DOI: 10.1101/2020.05.13.093146 (published DOI: 10.1111/tpj.14977)

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 knock-out lines whereas cslf9 knock-out lines had similar (1,3;1,4)-β-glucan content to WT. However, cslf9 mutants showed changes in the abundance of other cell wall-related monosaccharides compared to WT. Thousand grain weight (TGW), grain length, width and surface area were altered in cslf6 knock-outs and to a lesser extent TGW in cslf9 knock-outs. 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. ### Competing Interest Statement The authors have declared no competing interest.

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