Abstract
Fresh water cyanobacteria, such as Synechococcus elongatus PCC 7942 (hereafter refferd as S. 7942) and Synechocystis sp. PCC 6803, have multi-genome per cell. However, replication mechanism in fresh water cyanobacteria is largely unknown. In S. 7942, it has been predicted that replicational origin exists on the upstream region of dnaN gene (Liu and Tsinoremas, 1995, Gene, 172, 105-109), although experimental validation has not been carried out.
In order to investigate the site of the replicational origin in S. 7942, we constructed the assay strain S. 7942TK containing thymidine kinase gene (TK) in S. 7942 genome, since TK is essential for bromodeoxyuridine (BrdU) incorporation. The DNA replication in S. 7942TK was completely blocked by dark treatment. After addition of BrdU, DNA synthesis and BrdU labeling was initiated by light irradiation. DNA labeled with BrdU was isolated by immunoprecipitation, and analyzed for the newly replicated region using next-generation sequencer. The results indicated that the replicational origin exists on the upstream region of dnaN gene, consistent with previous report.
