Use of thiazole orange homodimer as an alternative to ethidium bromide for DNA detection in agarose gels

Abstract

Detection of polymerase chain reaction-amplified DNA fragments is commonly accomplished by visualizing the products in electrophoretic agarose beds with the use of ethidium bromide under ultraviolet light. However, ethidium bromide is mutagenic, and special handling and disposal precautions must be used. We report the use of a nonmutagenic dye, thiazole orange dimer (TOTO), which can be substituted for ethidium bromide. The excitation maximum for TOTO under ultraviolet light is 488 nm, and the absorption maximum is 510 nm, necessitating photographic filters different from those used for ethidium bromide for optimal results. Of particular importance in TOTO's use is the quantity used for each gel lane, since excess TOTO will cause unacceptable product mobility retardation. TOTO is only slightly more expensive than ethidium bromide. Overall, this stain provides very good visualization of polymerase chain reaction--amplified DNA bands in agarose gels. We believe the use of this safer reagent will become more widespread with increased regulation of laboratory activities.