I plan to have at least 10 ERT2-Cre activated and 10 control mice. I wonder with how big groups to start the injections, since I will inject repeatedly for 5 or 7 days. Some mice will likely drop out due to irritation by injection, some will drop out for other reasons. Some additional ones might drop out during behavioral analysis (mainly motor behavior and different cognition analyses, no additional invasive proceedures).
Thanks in advance for your input!
Hi Nadine,
From what I saw in our lab, this can actually vary from line to line - the effectiveness of the recombination is simply better is in some transgenic lines compared to others. On the other hand, we very rarely see any dropouts during the injection period - of course every now and then a mouse dies but if your injection technique is good exclusion due to injection is extremely rare.
The injection conditions are important though - tamoxifen or hydroxi-tamoxifen, how to dissolve, which dose, how often - so if someone already worked with your Cre-line, try to talk to them, find out what worked best for them. Or in case of doubt. run a couple of pilots.
To address your main question I would definitely analyse the recombination in every mouse after the behavioural tests. If you are testing cognitive abilities, you probably have a promoter that drives Cre expression in the brain. So isolate the brain and run an in situ on your target gene or a Cre immuno. You will then get an idea if you will have to exclude any mice from your behavioural analysis.
Ideally, you would have to test recombination in every batch of tested mice but this is not always feasible especially if you are targeting a small population of cells with a low expressing gene. If this is the case you especially need a robust and reproducible injection protocol that reliably gives you efficient recombination. Otherwise you will have a hard time deciding if a lack of phenotype in a mouse is due to individual variation or unsuccessful recombination.
I hope it helped,
Krisztina