Question

I'm trying to build some molecular gene switches into a model, and want to use both Cre and Flp. I understand how to invert gene segments with Cre, double floxing (e.g. with loxP and lox2272, like in this plasmid for example). To do the same with Flp/frt, I was looking at Plasmid constructs on Addgene, butmost results are only flanked by one (inverted) FRT site on each side. From my understanding, this would lead to flip-flopping back and forth, so when FLP is not expressed anymore, you'd get a 50/50 distribution. Can somebody explain to me why they do not use two frt sites, but instead one?

Answer 1

Cre and Flp recombinases work with loxP/lox2272 and frt sites, respectively, for DNA modification. While double floxing allows for controlled DNA conformations, single frt sites may lead to a 50/50 reversible configuration, depending on the experimental goals.

Step-by-step explanation:

Utilizing two distinct types of recombinase systems like Cre and Flp allows molecular gene switches to manipulate DNA sequences within cells. While using Cre recombinase involves double floxing with different lox sites (such as loxP and lox2272), Flp recombinase generally works with frt sites for DNA modification. Double floxing creates distinct DNA conformations that can be selectively reversed by Cre, whereas single frt sites can lead to a flip-flop situation, where Flp recombinase action can result in a 50/50 distribution of the inverted DNA segment when the Flp expression is turned off. The reason for not using two frt sites in some plasmid constructs could be related to the specific experimental goal or the need for a simpler recombination outcome compared to the more controlled outcomes possible with a double floxing strategy.