Project FAQ’s & Resources
1- Do you keep backup mice?
- We keep backup mice of either the same and/or previous generation for one month from the shipping date, in case more mice need to be generated.
- The mice can be kept for a longer time period if requested.
- We will inform you prior to sacrificing the backup mice, in case you need us to keep the mice longer.
2- What about my targeting vector and targeted ES cell clones?
- We are storing the positive ES cell clones from your project for one year at no charge. We will contact you after one year and you will have a choice to either receive the clones, or keep them in long term storage at our facility at extra charge.
- We also keep the targeting vector stored long term at no additional cost.
- You can request to receive these materials at any time.
3- What is the strain / substrain of the ES cells used for my mouse model?
|ES Strain||Substrain||Internal Name|
|FLP C57BL/6||C57BL/6 [N:J]||BF1|
|Hybrid (129 x C57BL/6)||129SvEv; C57BL/6N||BA1|
|FLP Hybrid (129 x C57BL/6)||129SvEv; C57BL/6N [N:J]||HF4|
Please note that all ES cell lines listed above were derived by ingenious.
4- What is the mouse strain that I can mate my mice to?
- Chimeras for all Hybrid, FLP Hybrid, C57BL/6 or FLP C57BL/6 ES cell strains are normally mated to C57BL/6N wildtype mice.
- Chimeras from projects using the 129 strain are mated to 129SvEv mice unless requested otherwise.
- Chimeras from projects using the Balb/C strain are mated to BALB/cAnN unless requested otherwise.
- These are the wildtype strains you should choose for further mating your mice.
5- What is the difference between somatic vs. germline Neo deleted mice?
- Vectors are designed with a selection cassette for antibiotic selection after electroporation and screening purpose. We commonly use the Neo selection cassette for most of our vectors. The Neo cassette is designed with flanking FRT recognition sites to be removed by FLP recombinase. It is highly recommended to remove the Neo cassette in the mouse model to avoid any interference of the Neo gene expression.
- One of the traditional methods to delete the FRT-flanked Neo selection cassette is via chimeras mating to FLP transgenic mice. The resulting mice (F1) will have the Neo deletion in somatic cells, which means in some cells Neo is absent and some cells still have Neo present.
- These mice will need to be mated to wild-type mice, and the offspring (F2) needs to be genotyped for absence of Neo to confirm that the Neo deletion has been transmitted in the germ cells.
- For projects that have vectors electroporated into our FLP ES cell lines, the FRT-flanked Neo selection cassette is deleted during expansion and culturing of ES clones. The chimeras are somatic Neo negative and are mated to wild-type mice to generate germline Neo negative mice (F1).
6- What does it mean that my mice are “FLP positive”? Do I need to remove the FLP transgene?
FLP recombinase is used to remove the FRT-flanked Neo selection cassette via chimera mating to FLP deleter mice or vector electroporating into FLP ES cell lines. Germline Neo negative mice will either retain or lose the FLP transgene. It is highly recommended to have the FLP transgene or any foreign transgene removed from your model before further breeding. To remove the FLP transgene, germline Neo deletion mice can be set up mating with wild-type mice.
Please refer to your genotyping document for detailed information on how to identify mice without the FLP transgene. Contact your project manager if you have questions.
7- How do I identify homozygous mice by genotyping?
After deletion of the Neo cassette, a footprint of the remaining FRT site or FRT/LoxP sites can be used for genotyping purpose. For most projects, primer set designed to detect Neo deletion can be used to identify homozygotes. For the specific primer set designed for your mouse model, please refer to your genotyping document for details on the genotyping strategy.
8- My gene is located on the X chromosome – now what?
If your gene of interest is located on the X chromosome, the F1 mice that were delivered to you are likely heterozygous female mice. These females can be set up mating to wild-type male mice to generate more heterozygous female mice and hemizygous male mice.
9- Additional Services at ingenious
We offer a range of additional services for our clients. You can choose from these services to supplement your efforts, or you can let us generate your experimental cohorts for you. Learn more about our Post Project Services including:
Post Project Consultation
- Sperm Cryopreservation
- Embryo Cryopreservation
- Generating Homozygous Mice
- Colony Scale Up
- Screening to identify homozygotes from heterozygote X heterozygote mating
10- Where can I find Cre-Expressing Mice?
We have compiled a list of Cre Mouse Respositories for your convenience.