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phenol/chloroform extraction of genomic DNA

genomic DNA phenol extraction

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6 replies to this topic

#1 Shifting Reality

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Posted 02 November 2013 - 09:02 AM

Hello everybody,

 

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?

 

Thanks,

 

SF

Attached Thumbnails

  • genomic DNA.jpg


#2 hobglobin

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Posted 02 November 2013 - 09:09 AM

looks more like a problem of the gel, therefore how does the size ladder look like?


Edited by hobglobin, 02 November 2013 - 09:10 AM.

One must presume that long and short arguments contribute to the same end. - Epicurus
...except casandra's that belong to the funniest, most interesting and imaginative (or over-imaginative?) ones, I suppose.

That is....if she posts at all.


#3 Shifting Reality

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Posted 02 November 2013 - 09:21 AM

Hello hobglobin,

 

It looks normal. It's an 1 % gel with 1 kb ladder.

 

http://kepfeltoltes....ltoltes.hu_.jpg



#4 hobglobin

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Posted 02 November 2013 - 10:40 AM

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.


One must presume that long and short arguments contribute to the same end. - Epicurus
...except casandra's that belong to the funniest, most interesting and imaginative (or over-imaginative?) ones, I suppose.

That is....if she posts at all.


#5 Shifting Reality

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Posted 03 November 2013 - 11:43 PM

Thank you very much ! ;)



#6 phage434

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Posted 04 November 2013 - 05:11 AM

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.



#7 mdfenko

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Posted 04 November 2013 - 05:24 AM

it could also be due to an incompletely dissolved pellet.


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