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Efficient generation of stable, heritable gene edits in wheat using CRISPR/Cas9

Author:
Howells, Rhian M, Craze, Melanie, Bowden, Sarah, Wallington, Emma J
Source:
BMC plant biology 2018 v.18 no.1 pp. 215
ISSN:
1471-2229
Subject:
Hordeum vulgare, Triticum aestivum, alleles, barley, diploidy, gene editing, heterozygosity, hexaploidy, inheritance (genetics), site-directed mutagenesis, wheat
Abstract:
BACKGROUND: The use of CRISPR/Cas9 systems could prove to be a valuable tool in crop research, providing the ability to fully knockout gene function in complex genomes or to precisely adjust gene function by knockout of individual alleles. RESULTS: We compare gene editing in hexaploid wheat (Triticum aestivum) with diploid barley (Hordeum vulgare), using a combination of single genome and tri-genome targeting. High efficiency gene editing, 11–17% for single genome targeted guides and 5% for tri-genome targeted guides, was achieved in wheat using stable Agrobacterium-mediated transformation. Gene editing in wheat was shown to be predominantly heterozygous, edits were inherited in a Mendelian fashion over multiple generations and no off-target effects were observed. Comparison of editing between the two species demonstrated that more stable, heritable edits were produced in wheat, whilst barley exhibited continued and somatic editing. CONCLUSION: Our work shows the potential to obtain stable edited transgene-free wheat lines in 36 weeks through only two generations and that targeted mutagenesis of individual homeologues within the wheat genome is achievable with a modest amount of effort, and without off-target mutations or the need for lengthy crossing strategies.