Mouse Genome Engineering Core

Transgenic Mouse

Transgenic Mouse

MGE core

  • Construct design expertise
  • Construct generation
  • DNA preparation
  • Pronuclear microinjection
  • Genotyping
  • Embryo transfer and generation of pups; tail clip
  • Transfer of founder transgenic mice and littermates as controls to PI laboratory
  • Advice on breeding strategies and strain maintenance

MGE Core or PI laboratory

  • Construct generation
  • Development of genotyping assay
  • Preparation of injection DNA
  • DNA preparation; Genotyping to identify transgenic founder animals
  • Analysis of transgenic mice

Transgenic Mice - the fine print…injection

  • DNA: MGE will obtain a minimum of 50µg of purified construct DNA from the Investigator initiating the project. Acceptable purification methods are Qiagen and sucrose gradient. MGE will also obtain proof of a valid genotyping assay for the construct. MGE will further obtain a statement from the Investigator with regard to any deleterious effects that expression of the transgene in question might have for mouse embryos and their development.

  • For one transgenic project, MGE will generate at least three transgenic founder animals (or transgenic embryos). To achieve this goal, MGE will strive to inject on average 200 embryos.

 

 

Responsibility for genotyping by 20 days of age, lies solely with the Investigator. MGE expects genotyping to be completed before the animals are weaned (20 days of age). Identified founder animals (together with a few negative animals as controls) are transferred to the Investigator. In case genotyping information is not available within a week of weaning, all animals will be transferred to the Investigator.

  • Should excessive embryo lethality – as defined by embryo death frequencies exceeding 30% for FVB embryos, and 40% for other strains - be encountered at the first microinjection experiment, MGE will postpone further injection experiments and request a new DNA preparation from the Investigator.
  • MGE's responsibilities are fulfilled when at least two founders have been produced, or when at least 50 viable pups (or embryos) have been produced, or when 300 embryos have been injected with a particular construct. In case there are no founders obtained from at least 50 pups or from 300 injected embryos, MGE will inform the Chair of the MGE Advisory Committee of it's efforts; at this stage it should be determined whether the biological effect of the construct (i.e. transgene expression during development) might be deleterious to the mouse embryo before further injection experiments are considered.

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