Fig. 4.

Effect of Cr(III) and Cr(VI) on synthesome-mediated and SV40 origin dependent in vitro DNA replication. In vitro DNA replication reactions were performed in the absence or presence of a range of concentrations of chromium chloride and sodium dichromate ion using a plasmid DNA template, (pBK-CMV), and affinity purified T-antigen, as described in the Methods. (A) Newly replicated DNA products were analyzed by electrophoresis through 1% neutral agarose gels following extraction and precipitation of the nascent DNA. Lane 1 shows the control reaction products formed in the absence of T-antigen. Replication reaction products include open circle DNA (oc-DNA), linear DNA (L-DNA) and supercoiled DNA (sc-DNA), and are shown in lanes 2–10. Reactions carried-out in the presence of sodium dichromate are shown in lanes 3–6; while reactions carried-out in the presence of chromium chloride are shown in lanes 7–10. The position of supercoiled DNA (scDNA); linear DNA (L-DNA); and open circle DNA (ocDNA) are noted to the left of the gel, as are the positions of higher order replication intermediates and the smaller Okazaki fragments formed during the reaction. (B) As described in the Methods, the amount of (³²P)-dCTP incorporated into in vitro DNA replication reaction products was quantified by scintillation counting. The values reported are the average of three independent experiments.