The study combined single-molecule fluorescence in situ hybridization (smFISH) and single-cell RNA sequencing (scRNA-seq) to map cell-type specific gene expression during early wheat spike development, profiling over 48,000 cells and identifying 21 spatial expression domains and 23 scRNA-seq clusters. Functional validation using induced mutants linked distinct expression patterns of LFY, SPL14, and FZP to specific meristematic regions, and imputed transcriptomes enabled the discovery of co‑expressed genes and gene networks governing spike formation.