DNA from cartilage?

Stuart Brown browns at ccu.umanitoba.ca
Fri Jul 9 16:08:45 EST 1993


In article <212hcn$57a at news.u.washington.edu> wolffie at u.washington.edu writes:
>
>I'm extracting DNA from cartilage for Southern blotting and PCR.
>Lots of glycoprotein or other such white glop coprecipitates with the DNA.  
>I don't know how much of a problem this is going to be, but I'd like to get  
>rid of the stuff if I can.
>
>The cartilage is minced, digested with proteinase K in 1% SDS-STE (sometimes  
>predigested with pronase or collagenase).  Then we add 1/3 vol of saturated  
>NaCl and extract this with Chloroform/isoamyl alcohol and precipitate with  
>isopropanol (we don't use phenol anymore).  The flocculent white precipitate  
>doesn't look like DNA and is much larger than one would expect; although it  
>does contain DNA, I expect this stuff is mostly digested glycoprotein  
>matrix.

>John Wolff,

	Try adding CTAB to 1%, NaCl to 0.7M, heat for 10' at 65C,
and then extract twice with chloroform.  Removes carbohydrates and
glycoproteins like a charm.  Ref. in Current Protocols (Red Book)
in section about preparing genimic DNA from bacteria.
	
	-Stu


-- 
Stuart M. Brown                             If you can remain cool when all 
U. of Manitoba, Dept. Plant Science         Around you are in panic,
Winnipeg, Manitoba, CANADA
browns at ccu.umanitoba.ca            Then you surely misunderstand the situation



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