Hello all,
This is somewhat embarrassing but I’m having difficulty genotyping my tamoxifen-inducible Advillin cre mice (B6.Cg-Tg(Avil-icre/ERT2)AJwo/J #026516) because the all the available protocols for this strain listed on Jackson’s website are for real-time PCR instead of traditional. I was wondering if anyone had a traditional PCR protocol they’ve used for this strain, or a generic iCre detection protocol.
Thanks,
Liz
Hey Liz,
Glad you got that mouse! Let us know how it is. Could be very powerful.
I don’t know how iCre differs from Cre in terms of sequence. But I would jus take their QPCR primers and do a regular PCR. The product is 150 bp. That’s enough.
I use Kapa use Kapa 2G mix, and it almost always works
https://www.sigmaaldrich.com/catalog/product/ROCHE/MGKITKB?lang=en®ion=US
Alternatively, why not do qPCR? Even for generic Cre now, I’ve moved to real-time. It’s faster and saves you the gel step.And if you really want, you can take the qPCR product and run it on a gel to convince yourself. I do Sybr qPCR. You don’t need to do the TaqMan that they do. I use Kapa Sybr.
Yeah, when I started out I didn’t genotype very often so it was cheaper to just run the gels. Now that I’m doing more PCR it might be worth moving to real-time.
I did try using their primers with a normal PCR protocol but I got like 6 bands in all samples, including neg control, and none of them were 150bp. I just thought I’d ask if anyone had something that they knew for sure would be successful before I went trying to troubleshoot.
Thanks!
Maybe just try the real-time. Sorry I don’t have anything better. Good luck!