I was genotyping some mice tail snips. It is a Cre Lox reporter mouse. The Cre is expressed using CRH promoter in the mouse brain and eGFP is the reporter gene. I isolated g-DNA and used it for PCR screening with primers supplied from Jackson labs. I checked a batch of 50 mice and found approximately 40 were CRH-Cre positive (both heterozygous and homozygous). These 40 mice were further checked for GFP expression and it turned out that some Cre positive mice were negative for expression of the GFP reporter gene. Is it possible for a mouse to be positive for Cre expression and yet be negative for reporter gene expression? Kindly let me know.
Hi Rashmi, it really depends on your breeding strategy (WT/Het/Homs of each transgene) and estimated probability of specified genotype offsprings.
Judging from your statement that "40 were CRH-Cre positive (both heterozygous and homozygous) ", I'm assuming your breeders were both Hets for Cre (in order to get you WT/Het/Hom Cre)...what about the second flox-eGFP transgene? Are you able to share more details on the mice strains for breeding?
In a nutshell, yes it is possible to have Cre positivity but flox-eGFP transgene offsprings if you'd perhaps crossed [Cre-hets X Cre-het:flox-eGFP hets] or [Cre-het:flox-eGFP hets X Cre-het:flox-eGFP hets].
Hope that helps!
Thanks Wei Wen Lim. I will look into the genotype of the breeder mice more closely
Hi Rashmi,
are you talking about protein expression of GFP is lacking, or are you trying to detect GFP "activation" by PCR? This clarification might help in answering your question better.
Pat