Previous work has established a genomic signature based on relative counts of the 16 possible dinucleotides. Until now, it has been generally accepted that the dinucleotide signature is characteristic of a genome and is relatively homogeneous across a genome. However, we found some local regions of the soybean genome with a signature differing widely from that of the rest of the genome. Those regions were mostly centromeric and pericentromeric, and enriched for repetitive sequences. We found that DNA binding energy also presented large-scale patterns across soybean chromosomes. These two patterns were helpful during assembly and quality control of soybean whole genome shotgun scaffold sequences into chromosome pseudomolecules.