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Visualizing DNA

Dyes for DNA Staining

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There are several different stains that can be used to visualize and photograph DNA after separation by gel electrophoresis. The following list explains some of the choices of stains, and the differences between them.

Ethidium Bromide

Ethidium bromide is likely the most well-known dye used for visualizing DNA. This dye can be used in the gel mixture, in the electrophoresis buffer or to stain the gel after it is run. Molecules of the dye adhere to DNA strands and fluoresce under UV light, showing where the bands are within the gel. Ethidium bromide is a potential carcinogen, so it must be handled with care. Because of the potential health risks of working with this gel, alternatives have been found.

SYBR Gold

SYBR Gold dye can be used to stain double or single-stranded DNA, or RNA. It was one of the first alternatives to ethidium bromide to hit the market and is considered more sensitive than ethidium bromide. The dye exhibits 1000-fold greater UV fluorescence once bound to nucleic acids. It penetrates thick and high % agarose gels and can be used in formaldehyde gels. Because fluorescence of the unbound molecule is so comparitively low, destaining is not required. Licence-holder Molecular Probes has also, since then, marketed SYBR Safe and SYBR Green as safer alternatives to ethidium bromide.

SYBR Green

The SYBR Green I and II stains, by Molecular Probes, are optimized for different purposes. Since they bind to DNA, they are still considered potential mutagens and should be handled with care. SYBR Green I is more sensitive for use with double-stranded DNA, while SYBR Green II is best for use with single-stranded DNA or RNA. Like ethidium bromide, these highly sensitive stains fluoresce under UV light.

SYBR Safe

SYBR Safe was designed to be a safer alternative to ethidium bromide and other SYBR stains. It is not considered a hazardous waste and can generally be disposed through the regular sewer systems (i.e. down the drain), because toxicity testing indicates no acute toxicity and little or no genotoxicity on Syrian Hamster Embryo (SHE) cells, human lymphocytes, mouse lymphoma cells and in the AMES test. The stain can be used with a blue-light transillluminator which causes less damage to the DNA being visualized and better efficiency for later cloning.

Eva Green

Eva Green is a green fluorescent dye that has been found to inhibit PCR to a lessor extent than other dyes, making it very useful for applications like quantitative real-time PCR. It is also a good choice when using low-melting point gels for recovery of DNA. It is very stable at high temperatures and has very low fluorescence on its own, but is highly fluorescent when bound to DNA. Eva Green has also been demonstrated to have very low or no cytotoxicity or mutagenicity.

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