Cytogenetics played an essential role in studies of chromosome structure, behavior, and evolution in numerous plant species. The advent of molecular cytogenetics combined with recent development of genomic resources has ushered in a new era of chromosome studies that have greatly advanced our knowledge of karyotypic diversity, genome and chromosome organization, and chromosomal evolution in legumes. This review summarizes some of the achievements of cytogenetic studies in legumes in the Phaseoloid clade, which includes several important legume crops such as common bean (Phaseolus vulgaris L.), cowpea [Vigna unguiculata (L.) Walp.], soybean [Glycine max (L.) Merr.], and pigeonpea [Cajanus cajan (L.) Huth]. In the Phaseoloid clade, karyotypes are mostly stable. There are, however, several species with extensive chromosomal changes. Fluorescence in situ hybridization has been useful to reveal chromosomal structure by physically mapping transposons, satellite repeats, ribosomal DNA genes, and bacterial artificial chromosome clones onto chromosomes. Polytene chromosomes, which are much longer than the mitotic chromosomes, have been successfully found and used in cytogenetic studies in some Phaseolus and Vigna species. Molecular cytogenetics will continue to be an important tool in legume genetics and genomics, and we discuss future applications of molecular cytogenetics to better understand chromosome and genome structure and evolution in legumes.