|Project initiation fee, non-refundable||TBD|
|Phase I: ES cell targeting phase: charged when targeted clones are identified or after 2 electroporations have been performed||TBD|
|Phase II: Production of chimeric mice, charged when chimeric mice are transferred to investigator||TBD|
Non-refundable fee covers assessment of DNA, provision of reagents, electroporation set up
Phase I: ES cell targeting
- DNA quantification and gel electrophoresis analysis
- The targeting construct is electroporated, ES cells are placed under drug selection and 200-250 drug-resistant colonies are picked
- Clones are provided to the investigator for DNA isolation and genotyping
- If no targeted clones are identified, a second electroporation will be performed and additional clones (up to a total of 480) will be provided for this same fee
- Projects are limited to 2 electroporations per construct
Phase I guarantees
- Based upon our continued success and a continual evaluation of ES cell stocks, we guarantee the efficiency of targeting of the ES cells and their germline transmission.
- Homologous recombination (targeting) is locus dependent. Therefore, we cannot guarantee that targeting will occur with a particular construct.
- Germline is not guaranteed for clones stored in 96 well plates for longer than 2 months. Why?
Phase II: Production of Chimeric mice
- Thaw, expansion and long-term storage of 1-5 ES cell clones
- Blastocyst microinjection of 1-4 targeted ES cell clones: initially, 1-2 clones are injected into approximately 24 blastocysts per clone. This is generally a sufficient number to generate at least 5 highly chimeric males per clone. If chimerism is low to moderate, another one or two targeted clones are injected.
Phase II guarantees
- Generation of 3-4 highly chimeric mice (>50% chimerism) from any of 1-3 targeted clones generated by the TTML using our standard laboratory ES cell stocks
- We cannot guarantee germline transmission of any one particular clone, however, germline transmission of the targeted ES cells is guaranteed
- Germline transmission is not guaranteed for other TTML ES cell stocks, including PRX-B6 and PRX-B6T, however, we guarantee that the parental lines are germline competent at the passage used for electroporation
Costs include the purchase of all mice used for the project; the housing of the surrogate females; and the housing of pups for up to 1 week post weaning.
- Additional clones can be expanded for a fee
- Housing costs: investigators will be charged for daily animal care if transfer of animals exceeds 1 week post weaning
- Use of other, non-routine strains for microinjection; includes the purchase and per diems of singly housed males for length of project