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ORF knockout question

biosci-request at net.bio.net biosci-request at net.bio.net
Wed Aug 11 11:09:41 EST 1999

In article <7oq6gn$7t1 at net.bio.net>,
 news at newsreader.wustl.edu, Alex Brands <abbrands at artsci.wustl.edu> wrote:
> Hello,
> I would like to generate a null allele of my new favorite gene in yeast.
> The gene I'm interested in is very close to the neighboring gene, and they
> are oriented in opposite directions, such that there is only 234 bases
> between the start codons. I'm concerned that the promoter of the
> neighboring gene extends into the coding sequence of the gene I want to
> knock out. If this is true, if I knock out the whole coding sequence, I
> will also disturb normal expression of the neighboring gene.
> With the tightly packed yeast genome, this must be a relatively common
> problem, how should I deal with it?
> Thanks for any input,
> Alex Brands

Why not knock out both ORFs and compare their phenotypes?  Is the second
gene essential? Also, you could make a smaller deletion of your gene of
interest, so as to start comfortably downstream of the ATG, and then check
to see if you've generated a null via Northern and/or Western.

Ed Davis, Ph.D.
ABL-Basic Research Program
Frederick, MD

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