- The composition and concentration of DNA loading dye can vary a lot. However, all DNA loading dyes contain two essential components:
- High-density reagent
- Tracking dye
- The simplest DNA loading dye contains at least one tracking dye and one high-density reagent. The most common and widely used DNA loading dye contains two tracking dyes and a high-density reagent.
- In addition to tracking dyes and high-density reagent, DNA loading dye can be supplemented with EDTA, SDS or Tris.
|Ingredients||Concentration(in 6x loading dye)||Remarks|
|High-density reagents: Usually a DNA loading dye contains one high-density reagent. Any of the following reagents can be added to the loading dye.|
|Glycerol||60% (v/v)||Interact with borate of TBE electrophoresis buffer. Do not use it with the TBE buffer.|
|Sucrose||40% (w/v)||Prone to mold-like growth|
|Ficoll 400||15% (w/v)||Can be stored at room temperature, quite stable|
|Tracking dyes: One or more tracking dyes can be added.|
|Bromophenol blue||Conc range: 0.03 – 0.5 % (w/v)Standard conc: 0.25% (w/v)||Migration corresponds to the migration of 4000 – 5000 bp long DNA fragments in 1% agarose gel|
|Xylene cyanol||Conc range: 0.03 – 0.5 % (w/v)Standard conc: 0.25% (w/v)||Migration corresponds to the migration of 4000 – 5000 bp long DNA fragments in 1% agarose gel|
|Orange G||Conc range: 0.15 – 0.5 % (w/v)Standard conc: 0.4% (w/v)||Migration corresponds to the migration of 40 – 45 bp long DNA fragments in 1% agarose gel|
|Optional/special purpose reagents|
|Tris.Cl (pH 7.6)||10 mM||Maintain the pH of loading dye|
|EDTA||60 – 100 mM||Protect DNA from DNA modifying enzymes by sequestering divalent cations|
|SDS||1%||Removes tertiary structure of DNA and eliminate protein from DNA|
- Sciarra & Elliott, 1960. A solubility study of the boric acid-glycerin complex. I. Solubility of boric acid in glycerin solution at 25 degrees. J Am Pharm Assoc Am Pharm Assoc. 49, 115-117. PMID-13854731; Full Text Links: wiley, readcube, booksc
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