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Using customized algorithms, the resulting data (ratio of each probe sequence replicated in early vs. late S-phase) can be converted into a form that can identify the genome.
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Description: This is a procedure for typing cells (cancer cells, stem cells, any kind of cells) based upon the order of replication of chromosome segments. In brief, cells from any source are pulse-labeled with 5-bromo-2deoxyuridine, sorted into early and late S-phase of the cell cycle by flow cytometry and the DNA replicated in each temporal compartment of S-phase is differentially labeled and hybridized to a DNA array consisting of evenly spaced probes from the entire genome. Using customized algorithms, the resulting data (ratio of each probe sequence replicated in early vs. late S-phase) can be converted into a form that can segment the genome and identify the order of replication of chromosome segments characteristic for a cell type. An alternative, if the cell line is difficult to label metabolically, is to sort cells into S-phase and G1-phase populations, hybridize differential labeled DNA from these sorted populations, and determine the ratio of each probe sequence in S vs G1. This provides similar data that can be evaluated by the same computational conversion. Advantages: More comprehensive (covers the entire genome) Less expensive (covers the entire genome for less than 1/20th what is needed for existing profiling methods) Much easier to interpret- the informative data for each cell line is distilled down to combinations of only about 1,000-2,000 segments of the genome that uniformly identify each cell type Measures very different properties of cells than any other method Focuses the analysis on the proliferating population of cells, which is particularly useful for stem cell and cancer technologies.