why does maxiprep always fail for this plasmid??

Mike Levin mlevin77 at mediaone.net
Mon Aug 20 21:07:17 EST 2001


Hi All -

   I'm hoping you can shed light on the following phenomenon. In my lab, we
often get plasmids in the mail. To use the construct, we transform them into
E. Coli (JM109 competent cells from Promega, mostly), grow them up in the
presence of an antibiotic (Ampicillin usually), and then do Quiagen
maxipreps to get the DNA. Usually it works fine - no problem. However, there
is one recent plasmid we got which just won't prep up! It transformed fine
onto an Amp plate, grew up fine in 300 ml of overnight shaking culture of LB
+ Amp, and when spun down made a bacterial pellet of the same size as others
done that day. However, when the Quiagen prep was done, the yield was very
low (< 0.1 mg/ml). The other 5 preps done that way worked fine. This
situation was repeated 3 times over the last month, and this one plasmid
just won't work! What could possibly account the fact that the bacteria grow
fine, but I can't get the plasmid prep out?? Any ideas would be greatly
appreciated - please email to mlevin at forsyth.org. Thanks!!

Mike Levin






More information about the Methods mailing list