To begin to explore cell cycle differences in gene expression as one possible cause of RPC heterogeneity, genes that had been identified in other settings as correlated with particular phases of the cell cycle were examined for variations in the single RPC profiles. These genes were divided into two groups, G1/S and G2/M, based upon their reported expression,Hupehenine which has been assayed primarily in cell culture. The heatmap shown in Figure 11A depicts a representative sample of the G1/S group of genes assembled from the literature. Genes such as PCNA, Rrm2 and the Mcms, whose protein products play important roles in DNA replication, were observed in a significant subset of RPCs. Rrm2, for example, was observed in 76% of the profiled RPCs. ISH on retinal cryosections confirmed that these genes were expressed in the ONBL. In the developing retina, the movement of RPCs is coordinated with the cell cycle so that mitosis occurs at the most scleral edge of the retina, just adjacent to the RPE, and S phase occurs toward the vitreal side of the ONBL. The two gap phases, G1 and G2, occur in the intervening space. Closer inspection of the section ISH patterns for Rrm2 and Mcm5 revealed that these genes are more strongly expressed toward the vitreal surface than the scleral surface, indicating they are predominantly detected in S phase cells. At E12.5, the expression pattern of these genes was more scattered Peimisine throughout the ONBL likely reflecting the observation that the precise migration patterns of RPCs with respect to the cell cycle do not occur at this early stage. DISH conducted for Rrm2 showed that 56% of Rrm2+ cells were -thymidine+ at E16.5 and at P0 this number increased to 61%. These data indicate that Rrm2 expression is enriched in S phase cells. However, the microarray data predicted a higher number of cells should express Rrm2 than was observed in the ISH experiments. These results most likely indicate that Rrm2 is expressed in both the G1 and S phases of the cell cycle, but at significantly higher levels in S phase, such that the detection by ISH picks up mostly S phase cells.This would be akin to the situation in serum starved and restimulated fibroblasts where Rrm2 was observed at lower, but detectable, levels during G1, with expression increasing significantly at the G1/S transition and into S phase.