TBE buffer precipitation
RMART...@biotechnet.com
s for years and
we frequently get precipitation forming. Does anyone know the cause of this
problem? How can you make up 10XTBE stock solutions that do not precipitate?
What problems can TBE precipitation cause if we use the buffer anyway in
our experiments (sequencing gels, agarose gels)?
Thanks in advance,
Rick in Indiana
Martin Leach
wrote:
Sometimes it does ppt. sometimes it doesnt (rare though). I use it when it
has ppt'd slightly and it works fine. A good tip to clean the ppt from the
bottle is to use glacial acetic acid ;)
Martin
--
..... Martin Leach Email:le...@mbcrr.harvard.edu
_|____ Dept. of Pharmacology Phone: (617) 638-5323
/ o / Boston Univ. School of Med. Fax: (617) 638-4329
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(BULLDOZER) \_ Boston MA 02118 "Not the old underpants on your
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fzes...@chip.ucdavis.edu
Concerning the ppt. of 10X TBE
My 10X TBE ppts. about a month or so after I make it. What I do is I just
stick the bottle in the microwave to warm it up and until the ppt. goes
away. I then just use it as normal and I don't have any problems for both
sequencing and agarose gels.
However in a previous lab to avoid ppt we made a 5X stock.
A 5X solution is would probably be better if the quantities you make of your
10X solution won't fit in your microwave.
Elizabeth
Klaus.M...@anu.edu.au
>
>
>
>s for years and
>we frequently get precipitation forming. Does anyone know the cause of this
>problem? How can you make up 10XTBE stock solutions that do not precipitate?
>What problems can TBE precipitation cause if we use the buffer anyway in
>our experiments (sequencing gels, agarose gels)?
>
>Thanks in advance,
>
>Rick in Indiana
The answer my friends seems to be blowing in the wind : DUST??
0.2 micron filter the 10xTBE and it stops precipitating (at least in my hands).
Micronucleation of salts is my conclusion and yes if the ppte forms, then
the sequencing gels don't get hot enough.
Cheers, Klaus
--------------------------------------------------------------------------
Klaus Matthaei
Gene Targeting
The John Curtin School of Medical Research
The Australian National University
Snail mail: Canberra, ACT 0200, Australia
E-mail: Klaus.M...@anu.edu.au
Landline: 61 6 249 3782
"I'd rather a full bottle in front of me than a full frontal lobotomy"
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Jim Owens
>we frequently get precipitation forming. Does anyone know the cause of
this
>problem? How can you make up 10XTBE stock solutions that do not
precipitate?
My guess is Tris carbonate is not very soluble in water. Where does the
Tris carbonate come from? The CO2 in air.
If I am correct, it is impossible to make 10X TBE that will not
precipitate sooner or later. When I make it, I filter it through a
0.45um millipore filter. This removes a slight haziness from the
solution and seems to retard formation of the precipitate when stored in
a bottle cleaned of any precipitate from the last batch of 10X TBE.
My 2!, for what it's worth,
Jim Owens
Mike
Klaus.M...@ANU.EDU.AU wrote:
>
> >Help! My lab has been making 10XTBE buffer solution
> >
> >
> >
> >s for years and
> >we frequently get precipitation forming. Does anyone know the cause of this
> >problem? How can you make up 10XTBE stock solutions that do not precipitate?
> >What problems can TBE precipitation cause if we use the buffer anyway in
> >our experiments (sequencing gels, agarose gels)?
> >
>
> 0.2 micron filter the 10xTBE and it stops precipitating (at least in my > > > hands). Micronucleation of salts is my conclusion and yes if the ppte forms, then the sequencing gels don't get hot enough.
>
Klaus, your suggestion certainly addresses the root of the problem.
However, you can go one step further and address particulates in your
glassware.
If you do a quick acid rinse of the bottle to filter into, a 20x TBE
solution will stay in solution for over a couple of months if handled
carefully. Even a 25x solution will work for a while. I suppose if you
could waste a large, plastic, all-in-one filtration unit, the results may
be even better.
Cheers,
Mike
the End
Filtration through Whatman paper sufficiently removes the dust
from your 10X TBE preventing precipitation. If stored in a old fashioned
bell jar with a stoppered outlet at the bottom you will prevent crystalized
material from falling back into the container and seeding the precipitate.
Jim
J. Graham
Jim Owens
jgr...@bronze.ucs.indiana.edu writes:
>If stored in a old fashioned
>bell jar with a stoppered outlet at the bottom you will prevent
crystalized
>material from falling back into the container and seeding the
precipitate.
I think this is very important. In my group I have not been able to
convince anyone but myself that the lip of the stock bottle needs to be
wiped off after pouring out 10X TBE.
Good luck,
Jim Owens