Mouse liver DNA?
U15767 at UICVM.UIC.EDU
Mon Dec 26 10:55:16 EST 1994
On Wed, 21 Dec 1994 09:00:46 -0300 you said:
>I am trying to make genomic DNA from mouse liver. Unfortunately, after
>prot K digestion of homogenized tissue and phenol/Chloroform
>extraction/ethanol precipitation, I am getting a flocculent white
>precipitate coming out with the DNA. I am not seeing the nice stringy
>genomic DNA in my precipitation that I expect. I have tried multiple
>phenol/chloroform extractions and precipitations but this white junk won't
>disappear. Any idea what this precipitate is? lipoprotein? what can be
>done to get rid of it? Maybe there is a better part of the mouse to make
>DNA from? I need lots for multiple southerns (15 5g/ southern).
>Any input would be greatly appreciated!
I have extracted a lot of DNA from many mouse livers, d it is probably the
easiest tissue to work with in terms of homogenization, extraction, etc. (At
least in controls..)) First, do you rinse or perfuse your livers to rid them
of excess blood? This is not a major concern, but might provide a cleaner
tissue sample. Second, after your phenol extraction, do you notice a thin
white layer separating your aqueous and organic phases? This is the
interphase and it could be loaded with lipid, depending on the age of your m
mice and/or the condition of their livers. It is important NOT to aspirate
this interphase with your aqueous layer. You don't mention how much DNA you
are trying to obtain, but with even only half a mouse liver you can get
quite a bit. Leave some aqueous phase behind and insure leaving the
interphase behind as well! If this is your white flaky stuff, you'll
eliminate it. I hope this helps!
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