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The eukaryotic genome has a hierarchicalthree-dimensional(3D)organization with functional implications for DNA replication,DNA repair,and transcriptional regulation.Over the past decade,scientists have endeavored to elucidate the spatial characteristics and functions of plant genome architecture using high-throughput chromatin conformation capturing technologies such as Hi-C,ChlA-PET,and HiChIP.Here,we systematically review current understanding of chromatin organization in plants at multiple scales.We also discuss the emerging opinions and concepts in 3D genome research,focusing on state-of-the-art 3D genome techniques,RNA-chromatin interactions,liquid-liquid phase separation,and dynamic chromatin alterations.We propose the application of single-cell/single-molecule multi-omics,multiway(DNA-DNA,DNA-RNA,and RNA-RNA interactions)chromatin conformation capturing methods,and proximity ligation-independent 3D genome-mapping technologies to explore chromatin organization structure and function in plants.Such methods could reveal the spatial interactions between trait-related SNPs and their target genes at various spatiotemporal resolutions,and elucidate the molecular mecha-nisms of the interactions among DNA elements,RNA molecules,and protein factors during the formation of key traits in plants.