Structural organization of alpha-satellite DNA in a single monkey chromosome

The organization of α-satellite sequences in a single monkey chromosome has been studied by restriction endonuclease analysis and molecular cloning. A somatic cell hybrid containing the monkey chromosome was isolated by cloning after fusion of the mouse L-cell line B82 (thymidine kinase minus) with primary African green monkey kidney cells and selective growth in HAT medium. Unlike the mouse cells, the hybrid cells contain DNA that hybridizes with the α-satellite DNA of the monkey. The presence of a single α-satellite containing monkey chromosome was demonstrated by Giemsa-11 staining and by the absence of both this chromosome and monkey α-satellite DNA sequences in cells after back-selection in bromodeoxyuridine. Hybridization of restriction endonuclease-digested hybrid cell DNA with a cloned segment of African green monkey α-satellite DNA showed distinctly different patterns from those observed with monkey total DNA. In particular, EcoRI and HaeIII restriction endonuclease sites are much more abundant in the satellite sequences in the thymidine kinase-carrying chromosome than they are in total satellite. A library of hybrid DNA was constructed in a λ bacteriophage. Analyses of purified recombinant phage that hybridized with α-satellite also indicated an abundance of EcoRI and HaeIII sites. Of nine phage studied in detail, no two showed identical distributions of the two restriction sites in the α-satellite sequences, suggesting the independent evolution of different domains within the single chromosome. These results indicate that the thymidine kinase-carrying chromosome contains distinct subsets (domains) of the α-satellite DNA of the whole monkey genome and further, that while the satellite sequence on the single chromosome is distinctive, it is also complex.