Identification of an ovuleless mutant and fine mapping of its causal Ol locus in cucumber (Cucumis sativus L.).

Ovule number sets the upper limit on seed yield in plants, yet the molecular control of ovule initiation remains largely unknown. Here, we characterised a spontaneous ovuleless (ol) mutant that bears round leaves, produces seed-empty fruits and completely lacks ovule primordia. Genetic analysis of 512 F₂ plants showed that the phenotype is governed by a single nuclear locus. Whole-genome resequencing of mutant and wild-type DNA bulks revealed a strong Euclidean-distance peak at the distal end of chromosome 2. Six newly developed InDel markers delimited ol to a 1.8 Mb interval, but suppressed recombination within this region prevented further reduction of the interval size. Leveraging the Tnt1 insertional background, we detected two retrotransposon insertions unique to the mutant: one in exon 8 of Csa2G377920, encoding a lectin receptor-like kinase, and the other in the promoter of Csa2G403160, encoding a DOG1-domain bZIP transcription factor. Quantitative RT-PCR showed that transcripts from both genes are nearly abolished in ol mutants. Spatial and temporal profiling indicated that Csa2G403160 is strongly expressed in female buds during ovule primordium initiation and is rapidly induced by the synthetic cytokinin CPPU, whereas Csa2G377920 exhibits weak, constitutive expression and is cytokinin-insensitive. Collectively, phenotypic, genetic and expression evidence pinpoint Csa2G403160 as the most likely causal gene underlying the ovuleless phenotype and highlight cytokinin-responsive bZIP signalling as a previously unrecognised layer in cucumber ovule development. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-025-01606-9.