phenol/chloroform extraction of genomic DNA - (Nov/02/2013 )
I have done phenol/chloroform genomic DNA extraction from cell cultures recently and I have problems about how the DNA looks after running on a gel. The samples look so knotty on the gel. (I enclosed a picture about that). Do somebody know what the problem could be? Are there too much contaminants in the sampes?
looks more like a problem of the gel, therefore how does the size ladder look like?
It looks normal. It's an 1 % gel with 1 kb ladder.
okay then there are several possibilities: excess salts, or carry over of ethanol or perhaps phenol/chloroform. Excess of salts you can remove with an additional washing step. And you should dry the DNA sufficiently long after washing and avoid carry overs during DNA extraction. A clue you can get sometimes by the smell of the sample tubes, i.e. ethanol or phenol you should detect this way usually.
Thank you very much ! ;)
I actually think the gel image looks pretty normal. Genomic DNA prepares long strands of DNA, and when loaded on a gel, the strands get mixed up and trapped by the gel. I think this is what is happening here. You could test this by mechanically shearing your DNA by pushing it through a thin hypodermic needle under pressure several times, which will shear the DNA. I'd predict that you will see this "problem" go away.
it could also be due to an incompletely dissolved pellet.