precipitation of oligos

[rupp at embl-heidelberg.de]

In article <UfeSrrS00Uh_I2YVpl at andrew.cmu.edu>, ab4u+ at andrew.cmu.edu (Anand K. Bachhawat) writes:
> Dear netters,
> 1) What is the best method to get 17-mer and 20-mer oligos in aqueous
> solution (that have been used for sequencing) back into a powder form?
> 2) What kind of recovery does one get when one precipitates 17-mer and
> 20-mer oligos using (a) ethanol and (b) isopropanol ? Are there any
> carriers that would help in getting a better recovery? 
> Thanks in advance
> Anand Bachhawat

Hi everybody,

a) why does nobody mention the easiest and fastest way, without any 
salts beeing involved : the n-butanol precipitation of oligos ??

You simply take 1 part oligo solution, add 9 parts n-butanol and vortex
it vigorously to homogenize the two liquid phases. Afterwards you spin the 
precipitate down using a tabletop centrifuge (15 minutes at high speed)
and discard the supernatant -- that's it !
!This even works straight from the ammonia solution!
The method was described by Sawadogo and VanDyke in NAR, Vol19, 3, p674
and is used by us in EMBL regulary for our in house synthesis service.

b) why do people still use PAA gels to check the purity of an oligo and 
NOT the much faster and less time consumtive way by running the oligos 
on a thin layer chromatography plate??
Take a silca gel 60 F254 plate (0.2mm) on aluminium (Merck), apply your
sample in 2 microliters of H2O and run the tlc plate in a tlc chamber
for 45 min using the following run buffer: 

	n-propanol             55% v/v
	conc. ammonia solution 35% v/v
	water                  10% v/v

This method is very sensitive (down to app. 50pmol of oligo will be seen)
and was described by Wu et al in "oligonucleotide synthesis: a practical
approach" IRL press, some years ago :-)).
It is used regulary in our lab as well !!!


Hope this helps,


Thomas RUPP