Phenol/Chloroform Extraction and Protease - (Sep/27/2005 )

Hi everyone,

I was wondering why a phenol/chlorform extraction is more efficient than just a phenol extraction.

My second question would be why is it better to use a mixture of proteases rather than using trypsin when hydrolyzing the peptidic bonds.
I'm thinking that mixture will provide enzymes which cut at different sites so the fragments are smaller that if you used trypsin.

Thanks

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hi

both phenol and choloroform do the job for separation of proteins and nucleic acid (see ambion's website or this topic)
in the ancient protocols, the extraction was done by a phenol step, followed by a chloroform step who was supposed to finish the separation AND remove the remaining phenol (because it was more hydrophobic)
Actually the mixture phenol chloroform IAA (25/24/1) is done in the same time to save time and to preserve sample (cause you loose a part of it when pick up back the upper phase).


that's what i think too.

fred

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