Identification of causal mutations in barley and wheat is hampered by their large genomes and suppressed recombination. To overcome these obstacles, we have developed MutChromSeq, a complexity reduction approach based on flow sorting and sequencing of mutant chromosomes, to identify induced mutations by comparison to parental chromosomes. We apply MutChromSeq to six mutants each of the barley Eceriferum-q gene and the wheat Pm2 genes. This approach unambiguously identified single candidate genes that were verified by Sanger sequencing of additional mutants. MutChromSeq enables reference-free forward genetics in barley and wheat, thus opening up their pan-genomes to functional genomics.

Department of Plant and Microbial Biology University of Zürich Zollikerstrasse 107 Zürich CH 8008 Switzerland

Institute of Experimental Botany Centre of the Region Haná for Biotechnological and Agricultural Research Šlechtitelů 31 Olomouc CZ 78371 Czech Republic

John Innes Centre Norwich Research Park Norwich NR4 7UH UK

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