isoamyl alcohol

Gys de Jongh GysdeJongh at planet.nl
Sat Mar 31 14:26:51 EST 2001


""Dr. Hiranya S. Roychowdhury"" <hroychow at nmsu.edu> wrote in message
news:3.0.6.32.20010327213024.00793b90 at cnmailsvr.nmsu.edu...
> OK ... call me a heretic -- and my comments blasphemous -- but there are a
> few "factual errors" in both the editions of "Maniatis". Amazingly, some of
> these are carried over from the first and into the second edition.
>
> I am not sure of the NaOAc - SDS observation. SDS is soluble and PDS is not
> ... this being the principle that led to the choice of KOAc over NaOAC in
> the classical alkaline lysis. Few Grad students (and even some beyond) ever
> actively think about this not so subtle implication. I use KOAC for
> everything and  sometimes I use AmmOAC. I used to use NaOAC several years
> back (as a Grad student) and have also used NaCl (Primarily for RNA work),
> but have never observed the phenomenon of NaOAC ppting. My Initial reaction
> ("Eh!") was because of that.
>
> Now, about the observation re: SDS-NaAc fluffy stuff: I believe that SDS
> pptd out due to local conc. increase. If you add 10 or 20% SDS directly to
> 2M NaOH while making a fresh bactrial lysis solution, same thing would be
> observed. To make the solution, one should add the reagents to water, or
> water to one of the reagents before adding the second. NaCl, having a far
> greater dielectric point over its acetate counterpart, will have little
> difficulty going back into solution. I doubt, therefore, that the fluffy
> ppt is a result of EtOH or 2-propanol, per se.

Thank you for the comments.
You are right.
Yesterday I tried combinations of : { NaAc , KAc , NaCl , KCl }
with SDS {10% , 5% , 1% }
with Ethanol { 60% , 70%}
After 1h at -20  _all_  are  _clear_  solutions.

Before the addition of etanol :
Some show precipitations right after the addition of salt . All precipitations
went back in solution at 60 degrees. Some were due to local high concentration
(NaAc , NaCl) . Some not : KAc and KCl gave stable prec. at rt.

After the addition of ethanol :
All solutions were clear.

After cooling for 1h at -20 degrees :
All solutions were still clear.

>
> BTW, I am at a loss about the 50ng/uL DNA in 10% SDS. I have to again do
> the "Eh!" for that. :-]

She was a student and added TE to her DNA pellet.
It foamed.
I asked her what she added and she said : "like we did yesterday , from the
little bottle with the aluminium lid". She looked very disappointed. I gave her
a hearty welcome to the core business of science : making more precise errors
every day , and decided to help her. So first I tried a few things with SDS and
lambda DNA.

>
> Kornelius' "tight precipitate" (with KCl and SDS) observation is explained
> above (SDS v PDS). This is independent of the conc. of either of the reagents.
>
> I am suffering from flu presently ...
I became a grandfather presently :)
I hope you are well again.

--
Gys









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