SYBR Green is a synthetic fluorescent dye used to quantify and visualize double-stranded DNA in molecular biology research. It is stable over a range of temperatures, binds non-covalently to DNA, and has a linear dynamic range of up to four orders of magnitude. SYBR Green is commonly used in real-time PCR and acrylamide gels, and is more sensitive than ethidium bromide. It can cause mutations and is a possible carcinogen, but is safer to work with and dispose of than ethidium bromide.
SYBR Green® is a fluorescent dye that binds to double-stranded DNA with high specificity. It is widely used in molecular biology research to quantify and visualize double-stranded DNA. SYBR Green® is a synthetic dye, introduced in the early 1990’s by Molecular Probes®, Inc., a subsidiary of Invitrogen&trade Corporation. It is an asymmetric cyanine dye, cyanines being a family of synthetic organic dyes used in photography, CD-R and DVD-R media, and a variety of molecular biology research applications.
Several characteristics make SYBR Green® particularly useful as a nucleic acid stain. It is stable over a range of temperatures, which means it can be used to monitor biochemical reactions that occur at high and low temperatures. It binds non-covalently to the DNA surface, meaning it will not interfere with the activity of most nucleases and DNA polymerases. It also binds to nucleic acids over a linear dynamic range of up to four orders of magnitude, so it can provide accurate DNA quantification over a wide range of concentrations.
The most common laboratory uses of SYBR Green® are the detection of double-stranded DNA in real-time polymerase chain reaction (PCR) and acrylamide gels. PCR is a technique for amplifying a few copies of DNA through repeated rounds of DNA replication. Real-time PCR measures amplification as it occurs, and since SYBR Green® binds to double-stranded DNA, measuring the fluorescence output of the DNA-dye complex allows the researcher to quantify the amount of DNA produced . As PCR cycles progress, there is an increase in fluorescence proportional to the amount of PCR product produced.
Acrylamide gels are ubiquitous in molecular biology laboratories, serving the commonly needed purpose of separating DNA fragments based on size. SYBR Green® allows the researcher to visualize DNA in an acrylamide gel by projecting light of the appropriate wavelength onto it. Other uses of SYBR Green® include the detection of nucleic acids in solution, in biochip applications and in flow cytometry. The DNA-dye complex absorbs blue light at 488 nm and emits green light at 522 nm.
SYBR Green® is very sensitive; it is 25 times more sensitive than ethidium bromide, another dye commonly used to visualize DNA. SYBR Green®’s high affinity for double-stranded DNA makes it useful for detecting low copy number DNA samples. It preferentially binds double-stranded DNA, but can also bind single-stranded DNA with reduced fluorescence. As with any DNA-binding molecule, SYBR Green® can cause mutations and is a possible carcinogen. It is, however, safer to work with and easier to dispose of than ethidium bromide.
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