Do you have a protocol for ethidium bromide staining in agarose gels?
For use in agarose gels:
- Add ethidium bromide to melted agarose to a final concentration of 0.5 µg/mL. Do not melt agarose that already contains ethidium bromide.
For staining agarose gels after electrophoresis:
- You can stain gels that have been run in the absence of ethidium bromide by covering the gel in 0.5 µg/mL ethidium bromide in water and gently agitating for 10 to 30 minutes.
- If necessary, gels can be destained by shaking in H2O for an additional 30 minutes. To stain RNA gels, you should minimize staining time and definitely include a destaining period.
What can ethidium bromide be used to detect and what is its sensitivity?
Ethidium bromide can be used to detect ssDNA, RNA, and dsDNA. This fluorescent dye intercalates between the stacked bases of nucleic acids, and exhibits an increased fluorescence at 590 nm. Ethidium bromide can detect down to ~1-10 ng/band of dsDNA in an agarose gel.
What amount of ethidium bromide do I need to stain nucleic acids in gels?
After electrophoresis, the gel should be stained in a 0.5-1.0 µg/mL solution of ethidium bromide in deionized water for 15 to 60 min depending on the thickness of the gel. As an optional step to reduce background fluorescence, the gel can be destained in deionized water for 15 to 30 min. Alternatively, ethidium bromide may be added directly to the agarose prior to casting. Add ethidium bromide to melted agarose to a final concentration of 0.5 µg/mL. This has the advantage of reducing the amount of ethidium bromide waste. However, this procedure may reduce the migration rate of nucleic acids.
引用和文献 (2)
引用和文献
Abstract
Immune-like phagocyte activity in the social amoeba.
Authors:Chen G, Zhuchenko O, Kuspa A,
Journal:Science
PubMed ID:17673666
Social amoebae feed on bacteria in the soil but aggregate when starved to form a migrating slug. We describe a previously unknown cell type in the social amoeba, which appears to provide detoxification and immune-like functions and which we term sentinel (S) cells. S cells were observed to engulf bacteria ... More
Cloning and pharmacological characterization of the guinea pig P2X7 receptor orthologue.
Authors:Fonfria E, Clay WC, Levy DS, Goodwin JA, Roman S, Smith GD, Condreay JP, Michel AD,
Journal:Br J Pharmacol
PubMed ID:18037910
BACKGROUND AND PURPOSE: The human, rat, and mouse P2X(7) receptors have been previously characterized, and in this study we report the cloning and pharmacological properties of the guinea pig orthologue. EXPERIMENTAL APPROACH: A cDNA encoding for the guinea pig P2X(7) receptor was isolated from a guinea pig brain library. The ... More