I wish to use phenol:chloroform extraction for the isolation of total RNA from murine spleen cells. The protocols I have read all include Sarkosyl in the denaturing buffer. Can this detergent be replaced with any other more common detergents? The extract will be used for downstream PCR reactions.
Thanks.
Interesting, the only RNA extraction protocols I have used do not include a detergent in denaturing buffer.
We used trizol/tri-reagent, but that is easily mimicked by adding guanidium thiocyanate into phenol chloroform.
And yes, I'm citing a 1987 paper :P http://www.ncbi.nlm.nih.gov/pubmed/2440339
The protocol is fairly easy, just don't breathe in too much of it/do it under a hood if necessary. Hope this helps as I don't have too much experience with sarkosyl lysis :(
I have just realised that the protocol(s) I have been looking at are an updated version this protocol. I am still not clear on the exact function of the Sarkosyl in the lysis buffer, but it seems that it may be unnecessary.
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