Replication of 5S ribosomal genes precedes the appearance of early nuclear replication complexes

The present work shows that replication of the 5 S ribosomal genes differs in time and 3'deoxyadenosine sensitivity from replication of other nuclear genes, in Allium cepa L. root meristems. Fluorescence in situ hybridization with the pTa794 DNA probe which contains a complete 410 bp 5 S gene from Triticum aestivum allowed to detect four clusters of 5 S genes in these diploid cells (2n = 16), two of them in the short arm of the smallest metacentric chromosomal pair 7. Replication of the 5 S ribosomal genes occurred very early in interphase, as discerned by their resolution as doubled spots only two hours after interphase was initiated in synchronous binucleate cells. Codetection of nuclear replication (by immunodetection of 5-bromo-2'-deoxyuridine incorporation) showed that the replication of the 5 S ribosomal genes occurred before any incorporation of 5-bromo-2'deoxyuridine could be detected in the nuclei. The earliest Br-DNA detected in these cells followed a radial pattern from different foci apparently dispersed along some chromosomal arms. These structures seem to represent early replication complexes, as a result of the displacement of multiple DNA forks from the foci known as pre-replication complexes where the replication machinery of the earliest replicating genes assembles. No consistent positional correlation existed between the formation of the early replication complexes and the already replicated 5 S ribosomal clusters. Finally, nuclear replication but not that of the 5 S genes was prevented by 3'deoxyadenosine, and the earliest replicating 5 S ribosomal gene cluster differed in both sister nuclei resulting from the segregation of one single chromosome in anaphase.