EvaGreen® dye is spectrally similar to FAM or SYBR® Green I, which means no change in optical settings is required for using an EvaGreen®-based master mix. However, there are several differences between the dyes.
Lower background fluorescence: EvaGreen® dye has less background than SYBR® Green I due to its novel “release-on-demand” DNA-binding mechanism.
Brighter and non-inhibitory: EvaGreen® is less inhibitory in the PCR reaction than SYBR® Green, permitting the use of saturating dye concentration for maximal signal and better high-resolution DNA melt analysis.
Safer and more environmentally friendly: EvaGreen® dye is the first and only PCR dye to date designed to be environmentally safe.
Dye stability: EvaGreen® dye is very stable both during storage and under PCR conditions. SYBR® Green I, on the other hand, is known to degrade following multiple freeze-thaw cycles and under PCR conditions. Moreover, decomposed SYBR® Green I is reported to be even more inhibitory to PCR.
Digital PCR: EvaGreen® dye is currently the only qPCR dye to be used in droplet digital PCR (ddPCR).
For most qPCR applications, we recommend using EvaGreen® Dye, 20X in water (31000), or one of our 2X EvaGreen® qPCR Master Mixes. EvaGreen® Dye, 2000X in DMSO (31019) is offered for specialized applications requiring a concentrated dye solution.
EvaGreen® Plus Dye is an improved alternative to Biotium’s original EvaGreen® Dye. EvaGreen® Plus Dye retains many of the essential benefits of the original patented EvaGreen® Dye, but has an improved signal-to-noise ratio that is more advantageous for a variety of DNA detection applications, including qPCR, digital PCR, and LAMP.
Lower background & higher signal compared to original EvaGreen® Dye
Better match for common qPCR instrument excitation sources
Compatible with qPCR & HRM®
Higher signal to noise offers advantages for digital PCR and isothermal applications
Directly visualize amplification products in gel
Improvements over the Original
Similar to our original EvaGreen® Dye, EvaGreen® Plus Dye is a green fluorescent nucleic acid dye that is essentially nonfluorescent by itself, but becomes highly fluorescent upon binding to dsDNA. EvaGreen® Plus Dye improves upon these properties with lower background fluorescence and increased brightness upon binding dsDNA. The dye has an excitation maximum that is a better fit for common qPCR instruments, resulting in improved sensitivity. As a result, EvaGreen® Plus Dye can give earlier Ct values and better signal discrimination. These properties offer potential advantages for digital PCR and isothermal amplification applications.
Why EvaGreen® Dye?
The unique properties of Biotium’s EvaGreen® dyes make them particularly useful in quantitative real-time PCR applications. Relative to other green dyes, such as SYBR® Green I, both EvaGreen® dyes are generally less inhibitory toward PCR and less likely to cause nonspecific amplification. Both EvaGreen® dyes enable direct visualization of the PCR product on a gel using a UV transilluminator or blue light box without the need for another gel stain. Furthermore, EvaGreen® dyes can be used at a much higher dye concentration, resulting in more robust PCR signal.
EvaGreen Dye and applications are covered by granted US and international patents. SYBR is a registered trademark of Thermo Fisher Scientific. HRM is a registered trademark of Idaho Technologies, Inc./BioFire Defense, LLC and its use may require a license.
Thiazole Green, which is structurally identical to SYBR® Green I Nucleic Acid Gel Stain, is one of the most sensitive stains available for detecting double-stranded DNA (dsDNA) in agarose, polyacrylamide gels and qPCR.