Nucleic Acids Research, Vol 24, Issue 21 4256-4262, Copyright © 1996 by Oxford University Press
F Buchholz, L Ringrose, PO Angrand, F Rossi and AF Stewart
Genomic manipulations using site-specific recombinases rely on their
applied characteristics in living systems. To understand their applied
properties so that they can be optimally deployed, we compared the
recombinases FLP and Cre in two assays. In both Escherichia coli and in
vitro, FLP shows a different temperature optimum than Cre. FLP is more
thermolabile, having an optimum near 30 degrees C and little detectable
activity above 39 degrees C. Cre is optimally efficient at 37 degrees C and
above. Consistent with FLP thermolability, recombination in a mammalian
cell line mediated by a ligand- regulated FLP-androgen receptor fusion
protein is more efficient at 35 degrees C than at higher temperatures. We
also document a mutation in a commercially available FLP plasmid (FLP-F70L)
which renders this recombinase even more thermolabile. The different
temperature optima of FLP, FLP-F70L and Cre influence their strategies of
usage. Our results recommend the use of Cre for applications in mice that
require efficient recombination. The thermolabilities of FLP and FLP-F70L
can be usefully exploited for gain of function and cell culture
applications.
ARTICLES
Different thermostabilities of FLP and Cre recombinases: implications for applied site-specific recombination
Gene Expression Program, European Molecular Biology Laboratory, Heidelberg, Germany.
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