Epigenetic Inhibitors Enhance Embryo-like Structure Induction in Wheat Anther Culture, but Green Plant Conversion Remains Genotype-Dependent

Wheat anther culture-mediated embryogenesis is strongly influenced by genotype responsiveness and remains limited by low embryo-like structure (ELS) conversion efficiency, frequent albino formation, and unstable developmental progression. In this study, the effects of DNA methylation inhibitors (5-azacytidine (AZC), 5-aza-2′-deoxycytidine (DEC), and zebularine (ZEB)) and the histone deacetylase inhibitor trichostatin A (TSA) on ELS induction, ELS conversion into green regenerants (GRs), albino frequency, and spontaneous genome doubling were evaluated in wheat anther cultures with contrasting embryogenic responsiveness. The results revealed strong genotype-dependent responses to treatments with epigenetic inhibitors. The spring wheat cultivar AC Abbey showed the highest overall anther culture response, whereas winter wheat genotypes displayed limited and variable ELS conversion competence. ELS formation was observed even in recalcitrant genotypes but was not consistently associated with efficient ELS conversion into GR. Multivariate analyses indicated stronger genotype differentiation at the conversion stage than during ELS induction, suggesting that this stage represents a major bottleneck in wheat anther culture-mediated embryogenesis. Among the tested compounds, AZC and TSA most effectively stimulated ELS induction, while AZC was associated with the lowest albino frequency. DEC showed the highest proportion of spontaneous doubled haploids. Overall, treatment with epigenetic inhibitors may enhance ELS induction but does not overcome genotype-dependent recalcitrance at the conversion stage under the applied anther culture conditions.