ES Cells-Based Gene Editing


Services

Knock-in/conditional knockout mice by gene targeting in ESCs

  •  This project is similar to the knock-in first with conditional knockout potential approach used by KOMP and EUCOMM, which allows the generation of knockout, reporter knock-in and floxed conditional knockout allele in one project. Unlike KOMP/EUCOMM's only choice of lacZ reporter gene, the core allows users to choose from several reporter genes such as GFP, nuclear GFP, ZsGreen, and tdTomato.
  • Bio-information analysis, targeting strategy and design/creation of reporter knock-in with conditional knockout potential targeting construct and other reagents.
  • Targeting by ES cell electroporation and microinjection to produce germline quality chimeras.
  • Breeding chimeras with C57BL/6 mice for germline transmission (GLT) to generate the heterozygous GFP reporter knock-in and conditional knockout mice. Design and test PCR genotyping methods, and genotype GLT F1 pups.
  • Guaranteed delivery for each project: one pair or more of heterozygous reporter knock-in with conditional knockout potential mice will be transferred to investigator.

Production of mouse chimeras by microinjection of KOMP/EUCOMM ESC clones

  • We will, after consultation, inject targeted ES cells provided from outside vendors (KOMP, EUCOMM, etc.) but we can not guarantee that these clones will produce chimeras, or that chimeras will go germline.
  • This service package includes expansion, cryopreservation, DNA preparation, and two microinjection sessions of up to three ESC clones per gene, including injection of a total of ≥100 embryos, and mouse housing until pups are 4 weeks of age.
  • Knock-in mouse production at Rosa26 or other designed locus by ESC targeting 
  • This is a guaranteed full-service to generate mice with insertions at Rosa26 locus or other designed locus. The insertion could be: Cre, CreGFP, CreERT2, GFPCreERT2, FLP and othercDNA.
  • Bio-information analysis, targeting strategy and design/creation of targeting construct and other reagents, design and testing of PCR genotyping methods.
  • Targeting by electroporation and microinjection of C57BL6-129S6 hybrid ESCs to produce germline quality chimeras.
  • Breeding chimeras with C57BL/6 for germline transmission (GLT) to generate the heterozygous Rosa26 knock-in mice. Genotype GLT F1 pups.
  • Guaranteed delivery for each project: one pair or more of heterozygous mice.

ESC clone expansion

  • The core will expand and freeze up to three ESC clones. The investigator will receive 2-4 vials/ESC clone and extracted DNA samples for verification of correct gene targeting.