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yeast zapping discussion

Tritech Research Tritech_Research at lamg.com
Mon Jul 8 11:25:53 EST 1996


Here is a little dialog about yeast electroporation.  If anyone else has
any comments, please jump right in!
Andrew
----------------------------
Dear Andrew,

regarding your e-mail from last week, the general consensus from the e-mails 
that I've recieved is that the smaller white colonies are almost certianly 
petites (rho-) colonies, that is yeast that have lost all or bits of their 
mitochondrial DNA and therefore are respiratory negative.  I also had a yeast

guy from Joe Sambrooks lab have a look at the plate and he agrees.  Another 
guy e-mailed the fact that electroporation is a great way to obtain petites.

Please  e-mail if you think this could be the explanation for your own work.

Gian Sberna.
--------------------
Well, it's something very interesting to consider, since I didn't know
that was a possibility.  I will look out for them when we do more
experiments.  However, it seems to me that the little one, just like
larger bright orange/pink yeasty looking colonies, and sometimes
slimy bacteria-looking colonies, come and go depending on how careful
the "sterile technique" is.

When do the little ones appear?  In our experiments, we always
do full voltage curves (since we are actually trying to study
yeast electroporation).  Transformants should peak at some
particular voltage, whereas contaminants are presumably
wild-type and will grow just as well on the plates with no zap
at all.

Do you have any data regarding this?

By the way, our yeast (strain M12b) seems to work at much
higher field strength than other stuff I've seen published.  Our
best yield of nice, big, healthy transformants is at about
1350 volts using a 1 mm gap.  Using a 2 mm gap yields
almost nothing.  Comments?

Andrew

TRITECH_RESEARCH at LAMG.COM




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