Complementary replication R- and G-band patterns induced by cell blocking at the R-band/G-band transition, a possible regulatory checkpoint within the S phase of the cell cycle.

Abstract

The characteristic band patterns of replication banding (dynamic banding) were analyzed. High-resolution (550-1,250 bands per haploid genome) G- and R-band patterns were obtained after 5-bromo-2'-deoxyuridine (BrdU) incorporation during early or late S phase. Thymidine-BrdU permutation culture methods, which arrest DNA synthesis at the R-band/G-band transition, allow complementary BrdU substitution. The RBI (R bands by BrdU using immunological staining) and GBI (G bands by BrdU using immunological staining) band patterns were complementary for all chromosomes. There was no overlapping, and every part of each chromosome was positively stained by one of the two banding procedures. Comparative analysis of RBG (R bands by BrdU using Giemsa staining) and RBI band patterns, as well as GBG (G bands by BrdU using Giemsa staining) and GBI band patterns, showed good congruency, displaying a very good band-for-band match. The congruency and complementarity found for these band patterns show that high concentrations of both thymidine and BrdU blocked S-phase progression near the R-band to G-band replication transition within the S phase. They also prove that BrdU incorporation is complementary and, therefore, demonstrate the existence of the R/G transition: a possible regulatory checkpoint within the S phase of the cell cycle.