Well obviously you would design primers homologous to the ERT2 part. That can of course differ between different mice based on the actual Cre-ERT2 construct that was used to make the transgenic mice.

I used these ones to detect VE-cadherin-CreERT2 when I had to cross these mice with other that contained my lox allele of interest, but also carried Emx1-Cre.

ERT2 fw 5'-G T G C C T G G C T A G A G A T C C T G

ERT2 rc 5'-A G A G A C T T C A G G G T G C T G G A

Of course it is also best to have a specific PCR for your constitutive Cre e.g. if you know at least the promotor sequence. Otherwise you can simply PCR for the Cre-ORF that should give a band for both Cre and Cre-ERT2 and compare that with your specific ERT2 PCR to sort out those mice crrying constitutive Cre and Cre-ERT2.

Uh oh. What happened Brandon? 

Thanks, Christian!  I will try that.

Bob, we received some mice from a lab that we thought were simply heterozygous for a floxed allele, which I confirmed with genotyping.  So I bred those mice to my cre of interest.  Once I did the genotyping, I knew something was wrong because I had too many cre+ mice (our primers detect ERcre and straight Cre's).  So I emailed the lab, and they said that it's likely that there was an ERcre in the animals that they gave us.  So I confirmed that there was a cre in the delivered animals (I should have done this first just to be safe).

So now I want to differentiate between ERcre and our Cre so I don't have to start over again with all cre- mice.

You could cross the original mouse to background (C57) and select Flox/+ Cre - mice and then interbreed littermates (Flox/+ X Flox/+) or Flox/+ X Cre; +/+. You would only be one generation behind, but clean.

Hi guys,

Has anybody tried CreERt2 primer sets for qPCR?! I am referring to primers bridging the interface of Cre and ERt2 you wrote about. I have a GFAP-CreERt2 mouse; hence want to verify transgenic expression exclusively in the brain but we are detecting endogenous Cre expression despite using Qiagen RNA extraction kit with DNase.

Thanks a lot!

Nadja

@Nadja Bjelopoljak did you find the primer? I am looking too, but I can find the right probe!

Thanks

Valeria

Usually we ordered a lot of transgenic mice from JAX, and they offer a lot of Cre and CreERT2 mice and in each webpage of specific cre, there is a PCR genotyping link which they provide the primers targeting either Cre or ER part. Below is an example of generic Cre, CMV-Cre, which is a germline Cre that's expressed in all tissues:

https://www.jax.org/strain/002471

And if you scroll down in this page you will find the link for genotyping:

https://www2.jax.org/protocolsdb/f?p=116:5:0::NO:5:P5_MASTER_PROTOCOL_ID,P5_JRS_CODE:22392,002471

Then you can go to IDT to order custom oligo for your PCR.