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. 2010 May 6;6(5):e1000939. doi: 10.1371/journal.pgen.1000939

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This is an open-access article distributed under the terms of the Creative Commons Public Domain declaration which stipulates that, once placed in the public domain, this work may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose.

This is an open-access article distributed under the terms of the Creative Commons Public Domain declaration, which stipulates that, once placed in the public domain, this work may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose.

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(A) A schematic showing the features of the oriII region. Two open reading frames, rctA and rctB (white arrows), border the region. The 11- and 12-mers (white or hatched arrowheads, respectively) are the putative RctB binding sites with GATC sequences (black dots). The origin also has a conserved sequence, 14-mer, and putative binding sites for DnaA (DnaA box) and IHF. (B) Electrophoretic mobility shift assay with fullymethylated DNA (Full), unmethylated DNA (None), the same DNA methylated in vitro by Dam (None + in vitro), hemimethylated DNA (Hemi) and the same DNA methylated in vitro by Dam (Hemi + in vitro). The Hemi DNA was methylated on the top strand. RctB amount was 0, 50 or 100 ng per 20 µl binding reaction. The free DNA band (black arrow head) refers to fragments not bound by RctB. The fraction of bound DNA was deduced from the loss of intensity of the free DNA band (Bound DNA = 1-free DNA). (C) Chromatin immunoprecipitation analysis of RctB binding in vivo. The precipitation was done with RctB antibody and the cells were either MG1655 (black bars) or its dam derivative (CVC1415, white bars), and each carried either the six-12mers (pGD61) or the empty vector (pRLM167). The histogram shows the average of three experiments.