Properties of nucleic acid staining dyes used in gel electrophoresis

Abstract

Nucleic acid staining dyes are used for detecting nucleic acids in electrophoresis gels. Historically, the most common dye used for gel staining is ethidium bromide, however due to its toxicity and mutagenicity other dyes that are safer to the user and the environment are preferred. This Short Communication details the properties of dyes now available and their sensitivity for detection of DNA and their ability to permeate the cell membrane. It was found that GelRed™ was the most sensitive and safest dye to use with UV light excitation, and both GelGreen™ and Diamond™ Nucleic Acid Dye were sensitive and the safer dyes using blue light excitation.

SYBR safe^TM efficiently replaces ethidium bromide in Aspergillus fumigatus gene disruption

“…Ethidium bromide is the most common stain used for nucleic acid detection on electrophoretic gels. This dye emits fluorescence when intercalated into DNA bases of nucleic acids and can interfere in both DNA and RNA synthesis (Singer et al, 1999;Ohta et al, 2001;Martineau et al, 2008;Haines et al, 2015). Ethidium bromide is classified as a strong mutagen and is genotoxic at the typical concentration for gel staining (Schagat and Hendricksen, 2013;Haines et al, 2015).…”

“…SYBR safe TM , a DNA stain developed by Invitrogen, is classified as non-genotoxic, non-mutagenic, and nonhazardous (Martineau et al, 2008;Evenson et al, 2012). This stain interacts with the DNA grooves instead of intercalating the DNA double stranded (Haines et al, 2015). Moreover, SYBR safe TM can be visualized under a blue light transilluminator, which does not damage the DNA (Martineau et al, 2008).…”

Optimization of Diamond Nucleic Acid Dye for Quantitative PCR

“…DD can penetrate the cell membrane, leading to interactions with genomic DNA (10); however, studies have shown that DD is less mutagenic and genotoxic when compared with EtBr (11). This reduction in toxicity may be due to the different binding mechanism of DD with DNA compared with other intercalating dyes, since DD does not bind between base pairs of DNA but instead binds externally (10). Currently, DD has not been evaluated for use in qPCR applications.…”

“…Diamond Nucleic Acid Dye (DD) is an external groove binding dye shown to have a similar sensitivity as SG and a limit of detection of 0.5 ng when detecting DNA in agarose gels (10). DD can penetrate the cell membrane, leading to interactions with genomic DNA (10); however, studies have shown that DD is less mutagenic and genotoxic when compared with EtBr (11).…”

“…Other fluorescent dyes that have been established for use in qPCR include EvaGreen (EG) (5), LCGreen (6), SYTO dyes (7), ResoLight (8), and BRYT Green (BG) (9), amongst others. One well-known issue with the use of SG in the laboratory is its mutagenic and toxic nature, as it can easily penetrate the cell membrane (10). SG is stated to cause mutations above the dosage of 33.3 g per plate in the Ames Test.…”