Oligo Percipitation

John Nash num208jn at MBDS.NRC.CA
Tue Jun 2 15:18:19 EST 1992


In article <1992Jun1.214109.3405 at bronze.ucs.indiana.edu>, jgraham at bronze.ucs.indiana.edu (the End) writes:
>Hello Bionetters,
>
>In my search for the simplest procedure for removing primers from a
>PCR reaction product, I was reminded that I had heard that you can
>recover the product by an amonium acetate/ ethanol percipitation and
>leave the primers behind. This supposedly worked well enough to
>sequence the products with an alternate primer directly.
>
>I would be most interested in hearing from anyone who has worked with 
>labeled oligonucleotides and various percipitation procedures.
>
>Thanks much,
>
>Jim
>jgraham at sunflower.bio.indiana.edu 
>
>Note: I'm amazed at how many people have responded to my questions
>with experiences with various commercial "kits". I hope this expensive
>trend does not continue, as it takes the procedure out of the users
>hands. It also has the effect of reducing the free flow of technical
>information between researchers. They might be nice to try but we do
>not use any commercially available kits or "mystery coumns" on a
>routine basis.
>

This is not a flame, nor a defence... let's call it a
counterviewpoint.

I use some kits and I have a good reason why.. I like glass milk, and
I like the way I can use it to clean up DNA from gels, after enzyme
treatments, etc, etc, but I am the only person in my group who does
**ANY** molecular biology, and I am not going to boil silica particles
in nitric acid to replace a $100 a year kit, and I don't have a TO or
grad student to do it for me ;-)   <== note smiley!

Sure... I make my own hyb solutions and the enzyme buffers which don't
come with enzymes, gel buffers, and the like... but then making and
testing ddNTP/dNTP mixes for T7 sequencing is a royal pain and so I
buy them.

I disagree with the ** NO KITS ** philosophy as much as I disagree
with the ** IT'S A KIT - LET'S BUY IT ** philosophy, and I think I'd
dismiss blanket statements that like they have "the  effect of
reducing the free flow of technical information between researchers"
as a gross generalisation... although the Taq polymerase story might
make me change my mind!

"Reasonable cost is interchangeable with reasonable timesaving" is
more like it!




..... but I write some of my own software ;-)


John Nash     (Internet:) Nash at biologysx.lan.nrc.ca.  
Email to my other addresses is forwarded automatically,
Disclaimer:  All opinions are mine, not NRC's!



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