Mouse Embryonic Stem (mES) Cells

Service Details

• NorCOMM ES Cell Withdrawal
• Non-NorCOMM ES Cell Line Expansion
• Non-NorCOMM ES Cell Line Antibiotic-Free Passage
• Generating Chimeric Mice from ES Cells

Service Charges

How To Order

Service Details

NorCOMM mES Cell Withdrawal

The CMMR is the repository and distribution centre for the NorCOMM mutant mES cell library. Before ordering your lines, we strongly recommend that you carefully review the data available for the CMHD and ESDB gene trap databases and blast the gene trap sequence tag against the Ensembl mouse genome, USCS genome browser and/or NCBI mouse genome to make sure that the sequence tag maps to your gene of interest and that it is in the sense orientation with the gene. It is also important to review the vector information to make the best choice among available lines. Upon receipt of a request, the bioinformatics associated with the requested line is reviewed to see if any obvious caveats are associated with a given line. If a problem upon bioinformatics review is detected, the Requester is contacted with the information and asked whether s/he would like to confirm or rescind the order. If no problems are detected, the Request is approved and put in the queue for expansion.

For each mES Cell Withdrawal lines are bioinformatically reviewed, thawed, expanded, frozen into aliquots and sequence validated by either 3'-RACE, RT-PCR, splinkerette PCR, inverse PCR or genomic PCR and sequencing of the resultant PCR product. SOPs associated with the procedures performed at CMMR can be found on our SOP website. Sequence confirmation of mES cell lines may be done at the CMMR or at the production facility (MFGC or CMHD). After sequence confirmation, two cryovials containing the expanded line are shipped on dry ice to the Requester at his/her expense. Requests normally take 6-8 weeks for expansion and validation before lines are shipped.

Not-for-profit investigators/institutions should carefully read and make sure they understand the CMMR Undertakings of Use. In order for their request to be processed, not-for-profit investigators/institutions must agree to abide by these terms.

For-profit investigators/institutions must execute a Material Transfer Agreement (MTA) with the mES cell production facility (University of Toronot for CMHD mES cell lines and University of Manitoba for MFGC mES cell lines). The CMMR will provide the appropriate template to the Requester. Upon return of two original signed copies of the MTA to the CMMR, the request will be put in the queue for expansion.

Service fees for mES Cell Withdrawal include bioinformatics review, line thawing, expansion, freezing and sequence validation. Service fees do not include shipping charges, including any costs to return dry shippers to the CMMR. Service fees must be paid in full by cheque or credit card, or by receipt of a valid institutional purchase order, before cells are shipped to the requester.

NorCOMM mES Cell Line Antibiotic-Free Passage

Due to the high-throughput nature of NorCOMM mES cell line production and medium-throughput nature of mES cell line distribution, pathogen testing is done in batches rather than testing of individual lines (see our pathogen testing policy for more information). While the CMMR is an approved mES cell facility for the generation of chimeric mice at the Toronto Centre for Phenogenomics Transgenic Core, eliminating the need for additional testing; other facilities may require that cells be tested for common murine pathogens prior to the generation of chimeric mice. Testing often needs to be preceded by antibiotic-free passage (AFP) of cells. The CMMR will produce AFP cell aliquots for a nominal fee in parallel with NorCOMM mES cell withdrawal when requested at the same time.

For each line, Antibiotic-Free Passage, involves three passages of mES cells without feeders and without antibiotics. Two vials of AFP cells are then supplied to the investigator with their expanded lines of interest as frozen cells in media or PBS, as requested. AFP ES cells are not suitable for experiments other than pathogen testing.

Service fees for Antibiotic-Free Passage include passage and freezing of cells. Service fees do not include shipping charges.

Non-NorCOMM ES Cell Line Expansion

Mouse ES cell line expansion is offered by the CMMR and TCP Transgenic Core as a service to the community to prepare mES cells for attempts to generate chimeras. Through the CMMR, investigators can recover mES cells from FIMRe member or other repositories for expansion and generation of chimeric mice. Please contact for more information.

Each mES Cell Line Expansion request includes line receipt, thawing, expansion, cryopreservation of new aliquots and growth in antibiotic-free media prior to mouse pathogen (IMPACT) testing. After growth in antibiotic-free media, cells are submitted to RADIL for testing. Testing for mouse pathogens is an important pre-screen of mES cells to assess the suitability of a line to generate chimeric mice.

Service fees include, receipt, thaw, expansion, re-freeze, and growth in antibiotic-free media prior to mouse pathogen testing.

Service fees do not include costs associated with withdrawal from other repositories, shipping or pathogen testing.

Generating Chimeric Mice from ES Cells

Through the TCP Transgenic Core, investigators can request attempts to generate chimeric mice from mES cells. These requests are normally preceded by NorCOMM mES Cell Withdrawal or Expansion of ES Cells Clones and Their Preparation for the Generation of Chimeras. When requested with an order for NorCOMM ES Cell Withdrawal, expanded mES cell lines can be thawed and transferred from the CMMR to the Transgenic Core for a nominal service fee. Through the CMMR, investigators can recover mice from mES cells supplied by FIMRe member or other repositories. Details regarding the generation of chimeric mice from mES cells at the TCP can be found on the Transgenic Services & Guidelines: Generation of Chimeras page. Please contact or for more information.

Service Charges

Service	TCP Investigators	Non-Profit Canadian Institution	Non-Profit International Institution	For Profit Institution
NorCOMM mES Cell Line Withdrawal, per line Includes bioinformatic review, line thawing, expansion, freezing and sequence validation Does not include shipping	$500	$500	$800*	$1250
NorCOMM mES Cell Line Antibiotic-Free Passage, per line Includes three passages of expanded line in antibiotic-free media and subsequent freezing of cells in vials for pathogen testing Available only in conjunction with NorCOMM ES Cell Line Withdrawal Does not include shipping or mouse pathogen testing	$100	$130	$160	$200
Additional Vial of NorCOMM mES Cells, per vial cIncludes expansion and freezing of additional vials of ES cells Available only in conjunction with NorCOMM ES Cell Line Withdrawal Does not include shipping	$160	$160	$256	$320
Genotyping Assay Development, per clone Includes primer design and development and testing of simplex PCR assays for wild type and mutant alleles of gene trap or gene targeted insertions at gene of interest If requested, primers and control DNA will be shipped to requester at requester's expense Does not include identification of genomic insertion site for gene trap clones or shipping	$375	$487.50	$600	$750
DNA Preparation for PCR, per sample Includes preparation of DNA from mouse tails or cells for use in PCR genotyping assays If requested, primers and control DNA will be shipped to requester at requester's expense Does not include identification of genomic insertion site for gene trap clones or shipping Note that these DNA preparations are NOT suitable for Southern blot analyses or other applications requiring double-stranded DNA	$2.25	$2.93	$3.60	$4.50
PCR Genotyping by PCR, per sample Includes simplex PCR assays for wild type and mutant alleles of gene trap or gene targeted insertions at gene of interest and genotype call If requested, primers and DNA will be shipped to requester at requester's expense Does not include identification of genomic insertion site for gene trap clones or shipping	$3.25	$4.23	$5.20	$6.50

*Local currency equivalent of 500 Euros; exchange rate equivalent to posted Bank of Canada rates at time of invoicing. For-profit investigators and institutions should contact the CMMR for pricing information.

How to Order

The Principal Investigator (PI) (and designate) must have an account on TCP LIMS. Please follow the Order Instructions for ES Cells.