There are a few possibilities:
The dye may have precipitated out of solution.
Heat the GeneGelRed solution to 45-50°C for two minutes and vortex to dissolve.
Store dye at room temperature to avoid precipitation.
If you are seeing high background staining of the gel, the agarose that you are using may be of low quality. Contaminants in the agarose may bind to the dye, resulting in increased background.
Yes. You can store precast GeneGelRed gels for up to a week. We recommend storing gels at room temperature in the dark. Do not store the gels at 4°C, because this will lead to dye precipitation and poor performance.
A 0.5 mL vial of GeneGelRed can be used to prepare 100 minigels (50 mL each) using the precast protocol, or for post-electrophoresis staining of 33 minigels in 50 mL staining solution per gel. Post-staining solution also can be re-used for staining 2 or more gels.
We do not recommend storing GeneGelRed in molten agarose for more than a few days.
GeneGelRed is a stable dye. You can use it in room light. However, we recommend storage of the dye in the dark between uses.
Yes. However, if the sensitivity decreases, use a fresh solution of the dye.
No, but de-staining with water can be performed if background is high.
We have tested a variety of DNA ladders from different suppliers with GeneGelRed with good results. Similarly, any common DNA loading buffers can be used. Band migration problems or smearing or smiling DNA bands are most commonly caused by overloading of ladder. If you are experiencing band migration problems with your ladder, please try reducing the amount of ladder you load. We recommend loading 50-200 ng ladder per lane for GeneGelRed precast gels.
No. We do not recommend reusing GeneGelRed gels after electrophoresis because the staining intensity can be decreased with sequential electrophoresis.
Yes, but it may be necessary to add some more dye to the re-melted gel for the best signal.
Yes. We recommend Qiagen or Zymoclean™ gel extraction kits or phenol-chloroform extraction to remove the dye from DNA.
GeneGelRed can be used to stain both ssDNA and RNA. Titration assays using a fluorescence microplate reader showed that the fluorescence signal of GeneGelRed bound to ssDNA and RNA is about half that of GeneGelRed bound to dsDNA.
GeneGelRed is ultra-sensitive dyes. Some users have reported being able to detect bands containing less than 0.1 ng DNA. However, the sensitivity of the staining will depend on the instrument capability and exposure settings.
GeneGelRed binds DNA exclusively by intercalation (http://link.springer.com/article/10.1007/s00249-014-0995-4).
GeneGelRed do not migrate through the gel as easily as ethidium bromide. It is not necessary to add additional dye to the running buffer, and the gel will be stained more homogenously than a gel stained with ethidium bromide.
GeneGelRed has been validated for Southern blotting. We recommend using the post-staining protocol for blotting applications.
Yes, customers have reported using GeneGelRed in glyoxal and formaldehyde agarose gels for precast staining of RNA.
Yes. Please use the post-staining procedure for DGGE and EMSA gels. For PFGE gels, the pre-cast or post-staining protocol may be used.
Yes, GeneGelRed can be used for Comet assay.
Yes, GeneGelRed is compatible with alkaline running buffer.
Customers have reported using GeneGelRed in cesium gradients. To extract GeneGelRed from DNA after cesium banding, we recommend adding SDS to a final concentration of 0.1% before butanol extraction. Alternatively, chloroform can be used instead of butanol for extraction.
Customers report good results using ZymoClean™ Gel DNA Recovery Kit from Zymo Research, GenElute™ Agarose Spin Column from Sigma, PureLink® Quick Gel Extraction kit from Life Technologies, illustra GFX PCR DNA and Gel Band Purification kit from GE Healthcare, High Pure PCR Product Purification Kit from Roche Applied Sciences, and GenJET gel extraction kit from Thermo Scientific.
Yes, see Ana. Biochem. doi: http://dx.doi.org/10.1016/j.ab.2012.09.003.
Yes, see Virol J. 2012, 9, 110.
Yes, use the post-staining protocol for polyacrylamide gels. For polyacrylamide gels containing 3.5-10% acrylamide, typical staining time is 30 minutes to 1 hour with gels of higher acrylamide content requiring longer staining time.,
Most of our products are stable at room temperature for many days, so in all likelihood the product will still work just fine. To be on the safe side, we recommend performing a small scale positive control experiment to confirm that the product still works for your application before processing a large number of samples or precious samples.