Mouse Modeling
The Embryonic Stem Cells and Mouse modelling (SCMM) facility runs a service of homologous recombination-based genome editing into Embryonic Stem Cells (ESC). The completion of the work will depend on the waiting list, the service works on a “first come first served” basis.
Skilled personnel provides investigators with a suite of tools and competence from the construction of the targeting vector(s) to breeding of chimeras for germline transmission, through intermediate steps that include generation of recombinant ESC clones and blastocyst microinjection of targeted ESCs. Rederivation of imported strains by embryo transfer is also run.
Each step-inquiry may be carried out independently from the others, but please note that animal holding, generation of transgenic strains and surgery/pharmacological treatments must be licensed by the Italian authority. Please login to intranet to download the mandatory documentation.
Construct design and generation
The strategy and the targeting constructs/vectors for the generation of the mutagenesis required (knock-out, conditionals, point mutation, targeted Cre or CreER deleters, gene replacement, etc…) will be agreed with the user.
Via cloning into vectors suitable for classical Homologous Recombination: Homology arms, suitable for PCR screenings, will be cloned into targeting vectors provided by the facility. Generation of the targeting vector will (may) require commercial BAC clones. This procedure should approximately take 1-2 months. | FEES: (Request Form) For IGB-ABT users: €3.000,00 For CNR (not IGB): €4.000,00 For external users: |
Generation of ESC clones for blastocyst injection
Via Homologous Recombination: The facility service includes the electroporation and isolation of heterozygous mutant ESC clones for the generation of transgenic mouse models. The fee includes the screening of 200 ES cell clones, screened by PCR and validated by Southern blot. For Southern blot analysis, the user has to provide suitable and tested probes, as agreed with the facility. The homologous recombination procedure should approximately take 2-3 months | FEES: (Request Form) For IGB-ABT users: €3.000,00 For CNR (not IGB): €4.500,00 For external users: €6.000,00 (+VAT) |
Microinjection
The injection fee is to be considered per number of injected blastocysts (n=80). The fee also covers animal purchase, cell culture costs and maintenance of up-to-six-week old chimeras. | FEES: (Request Form) For IGB-ABT users: For CNR (not IGB): €3.500,00 For external users:€5.000,00 (+VAT) |
Germline trasmission
The facility can provide breeding of chimeras for germline transmission. The service includes the purchase of wild type mice of the requested genetic background, housing, mating of chimeras, genotyping of the offspring. Germline transmission is expected within 4 months. | FEES: (Request Form) For CNR (not IGB): €2.000,00 For external users: |
Breeding, maintenance and genotyping of mutant mouse colonies
For the transgenic mouse lines generated by the facility, a service of breeding, maintenance and genotyping of mutant colonies is offered for the desired number of mice. | FEES: (Request Form) For IGB-ABT users: For CNR (not IGB): €0,90/cage/day For external users: €1,20/cage/day (+VAT) |
Strain Rederivation
The facility requires three males of proven fertility and genotype, which will be mated in our quarantine space with females of inbred/hybrid/outbred genetic background, according to each customer’s requirements. The service includes the purchase of embryo donor females, fosters, and maintenance of the offspring until weaning. Strain rederivation is expected in approximately 2 months. | FEES: (Request Form) For IGB-ABT internal users: For CNR (not IGB): €1.200,00 For any external user: €2.000,00 (+VAT) |
Generation of homozygous mutant ESC models
Via Homologous Recombination in ES cells: The homologous recombination procedure to generate homozygous ESCs should approximately take 6-8 months, and includes two rounds of targeting, each followed by removal of the selection cassette. For Southern blot analysis, the user has to provide suitable and tested probes, as agreed with the facility. | FEES: (Request Form) For IGB-ABT users: €6.000,00 For CNR (not IGB): €8.000,00 For external users: |
