Training & Education

CRISPR/Cas9 Training

Participants will learn to use CRISPR/Cas9 technology and will learn the scientific background needed to make genetically engineered mouse models, rat models or engineered mouse ES cells. You will learn both methods and the principles behind the technology. Research papers providing the experimental background underpinning methods used to engineer genes will be presented and discussed daily. A workshop on using software tools to acquire genomic sequence, Cas9 target selection, and design of DNA donor plasmids is included. You will learn all techniques necessary to produce and identify embryonic stem (ES) cells that have undergone homologous recombination with a targeting vector. Research papers providing the experimental background underpinning methods used to engineer gene targeted mice will be presented and discussed daily. Laboratory training includes daily laboratory sessions on molecular biology techniques such as DNA cloning, DNA sequencing, PCR, testing Cas9 ribonucleoprotein complexes for endonuclease activity, setting up a lab for ES cell culture, media and reagent preparation, normal ES cell culture, preparation of feeder cells, freezing and thawing of ES cells, identification of differentiated and undifferentiated ES cell morphologies, ES cell electroporation, picking ES cell clones, DNA preparation from ES cell clones in 96 well plates, and counting chromosomes in ES cell spreads.

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Embryonic Stem Cell Culture Training

Participants will learn all techniques necessary to produce and identify embryonic stem (ES) cells that have undergone homologous recombination with a targeting vector. Research papers providing the experimental background underpinning methods used to engineer gene targeted mice will be presented and discussed daily. Laboratory training includes setting up a lab for ES cell culture, media and reagent preparation, normal ES cell culture, preparation of feeder cells, freezing and thawing of ES cells, identification of differentiated and undifferentiated ES cell morphologies, ES cell electroporation, picking ES cell clones, DNA preparation from ES cell clones in 96 well plates, and counting chromosomes in ES cell spreads.

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Pronuclear Microinjection Training

Participants will learn all techniques necessary to produce transgenic mice. This includes superovulation, glass microinstrument fabrication, egg collection, identification of pseudopregnant females, surgical transfer of eggs to pseudopregnant recipients, and microinjection of DNA into the pronuclei of fertilized mouse eggs. A majority of trainees produce transgenic mice during training.

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Rat transgenic production training is also available.

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Blastocyst Microinjection

Participants will learn all techniques necessary to produce embryonic stem cell-mouse chimeras. This includes superovulation, glass microinstrument fabrication, blastocyst collection, identification of pseudopregnant females, surgical transfer of blastocysts to pseudopregnant recipients, and microinjection of ES cells into the blastocoels of C57BL/6 blastocysts. A minority of trainees produce chimeric mice during training.

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Mouse Embryo Transfer

Participants will learn all techniques necessary to transfer mouse embryos to the reproductive tract of recipients. This includes superovulation, isolation of pathogen free fertilized eggs from infected stocks, transfer pipette fabrication, identification of pseudopregnant females, surgical transfer of eggs to pseudopregnant recipients. All trainees produce offspring from transferred eggs during training.

Consult on design and construction of transgenes and targeting vectors; identification of transgenic founders; and breeding.