Grain number per spike (GNS) is a key factor influencing wheat grain yield and is determined by the combination of number of spikelet per spike and the number of grains per spikelet. This is associated with the progressive development of wheat inflorescence, an organ mixed of different meristem identities after flowering transition. Here, we explore the genetic regulation network governing wheat inflorescence development from a systematic view by integration of multi-omic data with population genetic approaches as well as functional validation by tilling mutant library. Single-cell transcriptomic and chromatin accessibility (sc-nuclei RNAseq and sc-ATACseq) data was generated from inflorescence tissue at important developmental stages including double ridge (DR), spikelet meristem initiation (SMI), glume primordium differentiation (GPD), floral meristem initiation (FMI) and terminal spikelet differentiation (TSD). In combination with the Stereo-seq, which captures the in-situ spatial transcriptomic profiling, we elucidate the transcriptome trajectory and chromatin dynamics during the progressive transition of different meristem identities, from shoot apical meristem (SAM) to inflorescence meristem (IM), then spikelet meristem (SM) and floral meristem (FM). Time-series analysis reveals cell-type and developmental stage-specific epigenomic dynamics concordant with transcriptome changes. Moreover, we built-up a core transcriptional regulation network (TRN) that likely driving the meristem cell identities transition. Integration TRN with genomic DNA variation and spike related phenotypes of collected wheat population, we have identified dozens of transcription factors including SPL, MADS, MYB, bHLH, YAB, TCP, NAC as candidate key regulatory factors for spike development. Furthermore, roles of TaYAB4/5, TaSPL6, TaMADS34, TaNAC50, TaMYB3 in regulation of spike development were confirmed genetically by tilling mutants and transgenic plants. The molecular regulation mechanism is also investigated. Collectively, we present invaluable resource and strategy for studying the mechanism underling wheat spike development in a systematic and high resolution manner.