Skip to main content.

Embryo Manipulation Services

The following embryo manipulation services are offered by the Transgenic and Gene Targeting Facility at the University of Kansas Medical Center. 

Click on a service to view details.

Responsibilities of the Investigator

  • Obtain IACUC approval for your project before requesting service.
  • Discuss your transgene design with the facility directors before initiation of the project.  Important: when designing your construct, ensure the transgene sequence is flanked by unique restriction enzyme sites to allow for easy removal of the vector backbone.  Isolation of your insert is essential, as inclusion of prokaryotic sequence may interfere with expression of your transgene.
  • Provide 25ug of circular plasmid DNA containing your construct at a concentration of 1mg/ml in TE buffer.
  • Provide a bacterial plate if submitting a BAC for injection.
  • Provide a restriction map and sequence of the construct including size, vector, promoter, gene of interest and/or reporter gene.
  • Submit a Request Form in iLabs to initiate your project.

Services Provided by the Facility

  • The Transgenic Facility will linearize, purify and quantitate your construct for injection.
  • We will use F1 hybrid FVB x C57Bl/6 mouse embryos unless requested by the user and approved by the facility.
  • Genotyping will be provided by the facility to identify transgenic founders.
  • The facility will guarantee two founders.  Exception: The facility does not prepare YAC DNA, and thus will guarantee only the injection and transfer of 200 embryos.
  • We cannot guarantee expression and/or transmission of your transgene.

In vivo Gene-Edited Mouse Production by CRISPR/Cas9

Responsibilities of the Investigator

  • Obtain IACUC approval for your project before requesting service.
  • Discuss your intended design and target sequence with the Facility Directors before initiation of the project.  The Facility does not currently perform gRNA or Cas9 RNA transcription or synthesis.  The Facility requests that RNAs to be injected be designed, tested, and produced according to rigorous validation procedures from approved vendors.  The Facility can assist in coordinating such efforts with a relevant Core that performs this work.  Please inform the Facility Directors if you intend to perform IVT in your lab or use a non-approved vendor.  
  • If your project design involves homology-directed repair, please inform the Facility and provide plasmid or a targeting oligo.  We recommend that targeting oligos are ordered from IDT.
  • Genotype the founders that result following injection of your CRISPR/Cas9 components.   NOTE: Please be aware that genotyping these deletions and/or mutations is complex and may take an extended period of time.   Your pups will be weaned onto your protocol and held in the Transgenic Facility for three additional weeks until your genotyping is complete, at which time the founders will be transferred to your animal room.  If your prefer, sequence-based genotyping can be performed by the Facility staff for an additional fee.
  • Submit a Request Form in iLabs to initiate your project.

Services Provided by the Facility

  • The Transgenic Facility will quantify and dilute your RNA (and DNA) components to the concentrations appropriate for injection.  
  • We will use F1 hybrid FVB x C57Bl/6 mouse embryos, unless requested by the user and approved by the Facility.The Facility will collect biopsies for genotyping.
  • The Facility will guarantee the generation of two founders for gene-deletion experiments only.  For HDR-based experiments, the Facility will guarantee injection and transfer of at least 250 embryos and/or 30 pups born, although reasonable efforts will be made to generate the mice requested.
  • NOTE: The Facility cannot guarantee germline transmission.
  • The Facility will provide tattoo identification of the resulting mice you choose to have transferred to your animal room.

Responsibilities of the Investigator

  • Obtain IACUC approval for your project before requesting service.
  • Discuss cell source with facility directors before beginning project. For requests to inject ES cells not provided by the Facility (including C57BL/6 cells from KOMP and EUCOMM), please contact Melissa Larson prior to obtaining those cells.  
  • Request karyotyping of your cells in iLabs prior to injection. 
  • Prepare cells the morning of injection and submit a single cell suspension on ice, or have the facility culture cells for you.
  • Develop a reliable genotyping assay (PCR, Southern, or both) for identifying positive mice from tail clippings or ear punches.
  • Testing for germline transmission is the responsibility of the investigator.
  • Submit a Request Form in iLabs to initiate your project.

Services Provided by the Facility

  • When requested, the facility will expand and prepare cells for injection.
  • Previously selected targeted ES clones will be microinjected into C57Bl/6 blastocysts, after which they will be implanted into pseudopregnant females.
  • We guarantee to inject and implant at least 60 blastocysts.
  • We cannot guarantee the production of chimeric mice, nor their germline potential.

Responsibilities of the Investigator

  • Have a minimum of five proven stud males available for each line.
  • If providing female mice, donors must be between three and four weeks of age, provided in groups of at least five.
  • Submit a Request Form in iLabs to initiate your project.

Services provided by the facility

Embryo cryopreservation will be accomplished by mating donor female mice to stud males of the line of interest. The facility can purchase wildtype females of either outbred or inbred background, or the investigator can provide the donor females for a reduced fee. (Required if cryopreserving homozygous mice.)

  • The facility will superovulate donor female mice and mate to investigator stud males.
  • Embryos will be harvested at d 3.5.
  • The facility will freeze morula/blastocyst stage embryos in straws with a controlled rate freezer.
  • Store straws in liquid nitrogen.
  • Thaw test straw to confirm viability of embryos.
  • We guarantee a minimum of 250 embryos frozen or three freezing sessions.

Responsibilities of the Investigator

  • Provide the facility with one or two males (2-4 months old) for each line.
  • Important: Each male must be of proven fertility prior to sperm freezing.
  • If you require that sperm from more than one male be frozen individually (rather than pooled), submit a requisition for each male as a separate line.
  • Although we request a 2-4 month old male, we can process any strain or age of mouse; however, it should be noted that age and mouse strain will affect the fertility of the recovered sperm.
  • Submit a Request Form in iLabs to initiate your project.

Services Provided by the Facility

The facility is employing the technique for sperm cryopreservation published by the Jackson Laboratory (2008):

  • Sacrifice male(s), harvest epididymides and collect sperm in cryoprotectant medium.
  • Sperm will be pooled from all males submitted for a line.
  • Freeze and store 15 straws in liquid nitrogen.
  • Thaw a straw to determine the motility of frozen-thawed sperm.

Responsibilities of the Investigator

  • Provide one male or 2 vials/straws of frozen sperm.
  • If providing a male for collection of fresh sperm, the male must be of proven fertility.
  • Submit a Request Form in iLabs to initiate your project.

Services Provided by the Facility

  • The facility will superovulate wildtype females of the requested strain.
  • Harvest oocytes and co-culture with sperm in vitro.
  • Transfer resulting two-cell embryos into pseudopregnant recipient females.
  • House recipient females and pups to weaning.

The Embryo Manipulation Core of the Transgenic and Gene Targeting Facility can accommodate special requests for service, including:

  • Embryo collection
  • Embryo injection
  • Sub-zonal lentivirus injection
  • Embryo bisection
  • Embryo transfer (mice and rats)
  • Vasectomy
  • Intrabursal injection
  • Artificial insemination

Talk to us about your project!  We will do our best to meet your embryo needs.  Fees will be determined on a per case basis and discussed in advance. Email: TGIF@kumc.edu with you project questions.

Transgenic and Gene Targeting Institutional Facility

University of Kansas Medical Center
3901 Rainbow Boulevard
G025 Hemenway
Kansas City, KS 66160
Fax: 913-588-2795
Phone: 913-588-7113
Email: TGIF@kumc.edu