CRISPR/Cas9-based genome editing technology has the potential to revolutionize agriculture, but many plant species and/or genotypes are recalcitrant to conventional transformation methods. Additionally, the long generation time of crop plants poses a significant obstacle to effective application of gene editing technology, as it takes a long time to produce modified homozygous genotypes. The haploid single-celled microspores are an attractive target for gene editing experiments, as they enable generation of homozygous doubled haploid mutants in one generation. Here, we describe optimized methods for genome editing of haploid wheat microspores and production of doubled haploid plants by microspore culture.
Keywords: CRISPR; Cas9; Doubled haploids; Genome editing; In vitro nuclease assay; Microspore culture; Microspores; Neon transfection system; Wheat; gRNA.