Nicking bromide

Fluorescence-based indicator reactions can also be coupled to measurements of enzymatic activity. For example, very sensitive assays of endonuclease activity can be performed using as a substrate complexes of double stranded DNA containing intercalated ethidium bromide, which is strongly fluorescent. On cleaving or nicking the DNA, the intercalated ethidium is released into the solution causing a pronounced decrease in fluorescence intensity. 

Addition of chloroquine or ethidium bromide in the running buffer resolves nicked DNA vs relaxed DNA (21). 

Electrophoresis in 1 percent agarose gels of nicked and closed circular PSMl DNA incubated with (a) cis- and (b) rran -[Pt(NH3)2Cl2] as function of time. After electrophoresis, gels were stained with ethidium bromide. Reproduced with permission from Reference 51. 

Fig. 5. Analysis of RNA complementary to MMTV DNA in 341 cells treated with (ADP-ribose) synthetase inhibitors. Total RNA from cells treated with 10 tM 3-ABm for 0 lanes a, f), 4 (lane b), 16 (lanes c, h), 32 (lane d), and 64 h (lane e), or with 10 mM 3-aminobenzoic acid for 16 h (lane g), was subjected to electrophoresis on 1.2% agarose gels, transferred to a nitrocellulose paper, and hybridized with nick-translated P]-labeled MMTV DNA specific for the env region. The size of the RNA were extrapolated from the migration of radiolabeled Hind III DNA standards (lane i) (4.4, 2.3, and 2.0) kilobase pair (Kbp) and 18S and 28S rRNA from the ethidium bromide staining pattern...

J In high concentration of ethidium bromide, the DNA can get nicked so it must be protected from the li t. 

FIGURE 7.4. Effect of intercalation on the sedimentation coefficient of X 174 replicative form, (a) Ethidium bromide (72% closed circles) (b) proflavin (67% closed circles) (c) actinomycin (75% closed circles). Open circles replicative form open triangles nicked DNA Hr dye bound/nucleotide. (From Ref. 10.)... 

---