Sister chromatid differentiation and isolabeling of chromosomes

Isolabeling observed during sister chromatid differentiation (SCD) was studied from human skin fibroblasts by the fluorescence-plus-Giemsa (FPG) technique. Bromodeoxyuridine (BrdU) was fed to exponentially dividing cells for 52 h to enable completion of two consecutive cycles of DNA replication. During this period, the late-replicating regions of some chromosomes were able to go through three replication cycles. These chromosome regions had evidently incorporated BrdU bifiliarly in both chromatids and hence, on staining with FPG, appeared isostained (isolabeled). Thus, incubation of exponentially dividing cells with BrdU for a period longer than that required for two cell cycles appears to be a suitable method for revealing the late-replicating regions of the genome, such as the X chromosome in a human female, as isolabeled. In another experiment with Indian muntjac chromosomes, isolabeled segments were darkly stained, which suggested unifilar incorporation of BrdU. In this case, unequal crossing-over or an unequal distribution of thymine residues probably is responsible for the isolabel.