Hi.
I have used Qiagen's plasmid maxi-preparation many times to amplify a
yeast genomic library, always with a good yield and no problems. The
library was in E. coli strain DH5-alpha.
Now, we are trying to amplify a single plasmid of this library that
contains a yeast gene we have cloned by complementation. The plasmid
was removed from yeast and transformed into E. coli strain BMH.
After 3 tries, we have been unable to get *any* yield whatsoever! I
have checked the buffers for pH. We made fresh lysis buffer [P2].
During the last run, samples were removed for the analytical gel and
will be analyzed. We are using the same exact kit and reagents that
have been used successafully before. We verified that the starting
colonies have the plasmid of interest.
The only thing I can think of is that there may be something different
about BMH strain that makes such a maxi-pred more difficult. Still,
getting no yield seems very strange.
If anyone has any ideas or suggestions, please share them.
Thanks very much!!
Louis Geller,
Graduate Student, Cell & Molecular Biology
Cal State Long Beach
lgeller at csulb.edu or bbbbd at sprynet.com
