PBR322 - maximizing yields

Paul N Hengen pnh at fcs260c2.ncifcrf.gov
Fri Aug 28 11:11:49 EST 1992


>bhjelle at carina.unm.edu () writes:

:Unfortunately, some things can only be done in pBR-based
:vectors and I have been plagued with totally miserable
:yields of one particular example of same for more than
:a month now.

I have two thoughts about your problem...

1. You may have cloned a segment of DNA that would prohibit
high copy number in any plasmid, in which case you will be
wasting much effort trying to amplify.

2. Why not spend some time subcloning into one of the
runaway plasmids that can be amplified simply by changing
temperature? This could save quite a bit of aggravation
and the bottle of chloramphenicol. See these refs...

B. E. Uhlin et al. Gene 6:91-106. (1979)
B. E. Uhlin et al. Gene 22:255-265. (1983)

Paul N. Hengen
National Cancer Institute
Frederick Cancer Research and Development Center
Frederick, Maryland 21702-1201 USA



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