The Facility offers a full set of gene targeting services, from advice on construct design to electroporation of a DNA construct into ES cells through the production of chimeric mice. A number of ES cell lines are available for targeting, including lines derived from the strains 129, C57BL/6J, C3H and B6D2F1/J. The Facility can also create chimeras from ES cells obtained from other sources, such as one of the gene trap consortia.
|Select and grow colonies (per 96 colonies)||$1000|
|Expand targeted ES cells (≤5 colonies)||$1000|
|ES Cell Microinjection|
|Prepare ES cells for microinjection (≤3 colonies)||$1000|
|Standard Microinjection of ES cells(C57BL/6 embryos)||$2000 + cost of mice and per diems|
|Non-standard Microinjection (non-C57BL/6 embryos)||$3000 + cost of mice and per diems|
|288 colonies picked||$3000|
|≤5 colonies expanded||$1000|
|≤3 colonies prepared for injection||$1000|
|Total||$8000 (+ cost of mice and per diems)|
Electroporation and Colony Selection. Your DNA construct will be electroporated into one of the available ES cell lines. Transfected colonies will be selected and expanded and split into 96-well plates. One plate will be frozen down; the other will be provided to you for DNA isolation and screening for targeted clones.
Note that the plates of frozen cells have a 6-month storage life. Delaying identification of targeted clones beyond six months risks loss of the clones.
Expansion of targeted clones. Once you have identified targeted clones, the Facility will expand up to five of these for you to re-test; additional colonies will be expanded at a fixed cost per colony. Expanded colonies will then be frozen for storage and future microinjection. We strongly recommend that you contact Dr. Barry Barnoski in the Cytogenetics Core and request that your ES cell clones be karyotyped prior to microinjection.ES Cell Microinjection
Cell preparation. Up to 3 ES cell colonies (targeted with the same construct) will be prepared for microinjection. At the investigator's request, additional colonies can be prepared at a fixed cost per colony.
Microinjection. Targeted clones will be injected into host embryos and the injected embryos will be transferred into pseudopregnant foster mothers. A minimum of 30 blastocysts/clone will be injected. Chimeric animals will be returned to the investigator for further breeding. At least 1 chimeric animal per construct will be provided.
Please note that we cannot promise delivery of germline chimeras. We strongly recommend that you contact Dr. Barry Barnoski in the Cytogenetics Core and request that your ES cell clones be karyotyped prior to microinjection.
Non-Standard Microinjection. This is the same service as standard microinjection, except that the recipient embryos are not C57BL/6. The costs are higher because working with these strains is more difficult. If you think you will require a non-standard microinjection, please contact the Facility Director first.
Again, please note that we cannot promise delivery of germline chimeras. We strongly recommend that you contact Dr. Barry Barnoski in the Cytogenetics Core and request that your ES cell clones be karyotyped prior to microinjection.
The Kimmel Cancer Center Web site, its content and programs, is provided for informational and educational purposes only and is not intended as medical advice, nor is it intended to create any physician-patient relationship. Please remember that this information should not substitute for a visit or consultation with a health care provider. The views or opinions expressed in the resources provided do not necessarily reflect those of Thomas Jefferson University, Thomas Jefferson University Hospital, or the Jefferson Health System or staff.
|Please send comments to: email@example.com. All contents copyright © 2002 Thomas Jefferson University. All rights reserved.|