oligo purification

Ed Rybicki ED at micro.uct.ac.za
Mon Mar 22 05:00:53 EST 1993


> I can't help but making an analogy to help clarify my position for all
> those who wrote in and advised to never check oligo's by gel as a waste of
> time.
>
> If you go out to buy a half a dozen eggs, do you open the lid to see if per

[omelette deleted]

> have had to do if you had purified?
>
> The answer is: YOU DON'T HAVE THE FOGGIEST IDEA, BECAUSE YOU'VE NEVER
> PURIFIED AN OLOGO AND DONE THE COMPARISON! But you still feel the need to
> write in and give lousy advice to someone who doesn't know better.
>

I didn't write in earlier, as I thought enough people would - but since
when is it a crime to offer comments based on good as opposed to bad
experience?  And yes, I have purified oligos made in-house, and yes, they
were a mixture of lengthmers, and YES, THEY WORKED FINE FOR PCR - of not-
particularly-well-conserved sequence(s) from a variety of plants (verified
by sequencing).

So - when you have a problem, check.  And if you are suspicious, check.
And if you are paranoid, check.  And use a program(me) such as anything
you can find if you must, but realise that someone once showed that
guesstimating oligos worked very nearly as well as using a programme, and
I found that if I had used any of the three best PC programmes available
at the time, I never would have synthesised the primers that I used
successfully to amplify the complete range of cereal geminiviruses,
because they were "unsuitable" - and I couldn't have used any other
sequences because the consensus was sufficiently high for oligo making
only in the UNSUITABLE sequences I used.

So - don't be put off from just trying 'em out anyway, un-computer
predicted, and unpurified!  Having said that, I've just advised someone
with a problem to purify her oligo, because the quality control wasn't
what it should have been, but we haven't wasted any money yet!

  ____________________________________________________________________
 | Ed Rybicki, PhD             |     "Emancipate yourselves from      |
 | (ed at micro.uct.ac.za)        |            mental slavery            |
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