Why is my Agarose gel glowing?
Posted 23 September 2004 - 11:40 PM
Can anyone suggest me why is this happening and how to get uniform stained gel?
Any help would be highly appreciated.
Posted 24 September 2004 - 04:06 AM
in summary: you can stain your gel after the run, or you can lower the concentration of etbr in you gel and the effect will disappear or become less noticable, respectively.
Posted 24 September 2004 - 06:21 AM
Posted 24 September 2004 - 06:51 AM
Posted 24 September 2004 - 08:18 AM
But while viewing the gel in UV transilluminator, I find the gel fluoresce gradually from bright to light.
Which way the variation goes, from side to side or from end to end? You may also check your light box to make sure that the bulbs light evenly.
Posted 26 September 2004 - 03:25 AM
I think probably it's due to the amount of EtBr you loaded into the gelaas I have this problem before. The DNA is negative charged while EtBr is positive charged, that's why it seems moving towards the wells under the UV transilumination. I usually use the pipette tip's end to slightly touch the EtBr solution and that's enough to prepare 100ml gel slab.
Posted 25 October 2004 - 11:43 AM
Posted 26 October 2004 - 01:53 PM
Another problem that may be influencing the photograph is the fading of bands on the gel, this is due to the DNA being fragmented by the UV, and there is not a lot that can be done about it... my only suggestion is to keep the gel exposure time to a minimum, especially if you are trying to extract the bands for further analysis.
Posted 26 October 2004 - 03:04 PM
Also, I notice that this gradient pattern usually becomes more frequent the longer you wait for the agarose to cool down before adding EtBr. I usually add my EtBr to my melted agarose when it is about 60 C...then I run away out of the lab before I breathe in any EtBr fumes. I don't know if there are such fumes, but I'm always suspicious of these things.
Posted 10 November 2004 - 08:46 AM
Posted 11 November 2004 - 06:51 AM
the best method to stain ur gel with et br willbe to strain it after the gel run is over. also u can try using lesser concentrations of etbr and stan them for a small duration to get sharp view of the bands. erbr will move through the gel if u are loading it along with the along with the buffer.