The Facility can perform a PCR assay to test for the presence of over 60 species of mycoplasma in cell culture. 

Investigator should provide the facility with cells in a 1.5 ml tube: 

• Suspension cells: count 10⁴ - 10⁵ cells and resuspend in 1 ml PBS or media 
• Adherent cells: scrape cells from one well of a 6-well plate and resuspend in 1 ml of PBS or media
  •  Do not dissociate cells with trypsin or EDTA; these agents disrupt mycoplasma. 

Cell suspension should be frozen and stored at -80. Alternatively, Investigators may provide the Facility with a vial of frozen cells or cells in culture.

Responsibilities of the Investigator

• Provide the facility with a vial of frozen cells or a plate of growing cells.
• Provide a protocol for culture.
• Submit a Request Form to initiate your project.

Services Provided by the Facility

• The facility will culture your cells in the appropriate medium, expand and freeze several aliquots.
• Frozen cells will be maintained in the facility's auto-fill liquid nitrogen storage tank, unless requested otherwise by the investigator.

The Facility will conduct optimization of electroporation parameters for your cells with our Neon NxT Transfection System.

The Transgenic Facility provides an initial consultation to all investigators wishing to create a gene-modified mouse or cell line. After the consultation, reagents can be designed and validated. CRISPR reagent validation includes reagent design, reagent validation in embryos or a relevant cell line, and optimization of the PCR reaction for detecting mutations at the target site. For more information, refer to the Fee Schedule or send us an email at tgif@kumc.edu.

The Facility employs CRISPR-Cas9 technology to generate gene edits in both iPSC and cancer cell lines.

• The Facility will design and order CRISPR reagents (crRNA from IDT) and validate activity in your cells.
• After an active guide or guides have been identified, the Facility will design a targeting construct for knock in projects.
• The Facility will electroporate RNP with or without a repair template for the production of gene-modified cells.
• The Facility will genotype all clones resulting from CRISPR-Cas9 electroporation to identify those clones possessing the mutation of interest.

The Transgenic and Gene Targeting Facility can derive primary human fibroblasts from biopsy/autopsy tissues.

Responsibilities of the Investigator

• Provide the facility with living tissue in DMEM/RPMI at room temperature or on ice.
• Submit a Request Form in iLabs to initiate your project.

Services Provided by the Facility

• The facility will derive primary fibroblasts from the tissue in the appropriate medium and freeze several aliquots.
• Frozen cells will be maintained in the facility's auto-fill liquid nitrogen storage tank, unless requested otherwise by the investigator.

The Transgenic and Gene Targeting Institutional Facility can reprogram primary fibroblasts into iPS cells.

The Transgenic Facility can differentiate iPS cells into different somatic cell types. Reach out to tgif@kumc.edu to discuss your cell needs.

The Facility can develop an assortment of unguided or guided organoids from iPSC.  Reach out to tgif@kumc.edu for more information.

The Facility can provide CRISPR-Cas9 gene-targeting services in mouse ES cells.