Novel semi-dwarfing alleles in wheat generated by TILLING in GA20ox1

The plant hormone gibberellin (GA) is involved in a wide range of developmental processes in plants, notably elongation growth. Semi-dwarf mutants involving GA biosynthetic and signalling components were important contributors to the Green Revolution in rice and wheat, respectively. To develop novel semi-dwarfing alleles in wheat we have screened an EMS-mutagenised population in the UK spring cultivar Cadenza for mutations in components of the GA pathway. RNAseq data from elongating stem tissues of wheat revealed transcripts from several GA20ox paralogues including GA20ox1. Using a range of TILLING methods including CelI cleavage and high-resolution melting we identified candidate loss-of-function alleles in the three homoeologues of GA20ox1. Mis-sense alleles were tested for effects on enzyme function by heterologous expression in E. coli. Mutations confirmed as full knock-out alleles were combined by crossing, yielding a triple homozygous mutant that was 12cm (14%) shorter than wild-type plants. Preliminary analysis of the segregation of the progeny of triple heterozygote plants (AaBbDd) suggests a gene dosage effect with each wild-type allele contributing approximately 2cm to plant final height.