ANALYSIS OF REPLICATION TIMING PROPERTIES OF HUMAN X-CHROMOSOMAL LOCIBY FLUORESCENCE IN-SITU HYBRIDIZATION

We have used fluorescence in situ hybridization on interphase nuclei o f normal female cells to compare the replication timing patterns of ge nes on the human X chromosome that are known to escape X inactivation with those that are inactivated. By this procedure it was possible not only to determine the relative time of replication of the earlier-rep licating allele for different loci but also to estimate the degree of asynchrony of replication of the two alleles for each individual locus . Loci such as HPRT and FRAXA, which are normally inactivated, display ed a high degree of replication asynchrony, whereas loci that are not inactivated (ZFX and RPS4X) were found to replicate very synchronously . Interestingly, examination of XIST, which if expressed only from the inactive X chromosome, by this procedure revealed that it also replic ated asynchronously, with the expressed copy apparently replicating fi rst. Therefore, by examining different loci from the X chromosome it w as determined that there is a strict correlation between the expressio n and relative time of replication of individual genes.