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Updated: May 29, 2026
Derivation of Stem Cell Lines from Mouse Preimplantation Embryos
Published on: August 20, 2017
Optimization of protocols for derivation of mouse embryonic stem cell lines from refractory strains, including the
Timothy J Davies1, Paul J Fairchild
1Sir William Dunn School of Pathology, University of Oxford, Oxford, United Kingdom.
Abstract:
The derivation of pluripotent embryonic stem cells (ESCs) from a variety of genetic backgrounds remains a desirable objective in the generation of mice functionally deficient in genes of interest and the modeling of human disease. Nevertheless, disparity in the ease with which different strains of mice yield ESC lines has long been acknowledged. Indeed, the generation of bona fide ESCs from the non obese diabetic (NOD) mouse, a well-characterized model of human type I diabetes, has historically proved especially difficult to achieve. Here, we report the development of protocols for the derivation of novel ESC lines from C57Bl/6 mice based on the combined use of high concentrations of leukemia inhibitory factor and serum-replacement, which is equally applicable to fresh and cryo-preserved embryos. Further, we demonstrate the success of this approach using Balb/K and CBA/Ca mice, widely considered to be refractory strains. CBA/Ca ESCs contributed to the somatic germ layers of chimeras and displayed a very high competence at germline transmission. Importantly, we were able to use the same protocol for the derivation of ESC lines from nonpermissive NOD mice. These ESCs displayed a normal karyotype that was robustly stable during long-term culture, were capable of forming teratomas in vivo and germline competent chimeras after injection into recipient blastocysts. Further, these novel ESC lines efficiently formed embryoid bodies in vitro and could be directed in their differentiation along the dendritic cell lineage, thus illustrating their potential application to the generation of cell types of relevance to the pathogenesis of type I diabetes.
Insights
Researchers developed a new method to derive embryonic stem cells (ESCs) from difficult mouse strains, including the non obese diabetic (NOD) mouse. This breakthrough facilitates disease modeling and genetic studies in diverse mouse models.
Area of Science:
- Stem Cell Biology
- Developmental Biology
- Genetics
Background:
- Deriving embryonic stem cells (ESCs) from diverse mouse genetic backgrounds is crucial for disease modeling and functional genomics.
- Generating bona fide ESCs from certain mouse strains, like the non obese diabetic (NOD) strain, has been historically challenging.
Purpose of the Study:
- To develop and optimize protocols for deriving ESCs from genetically challenging mouse strains.
- To establish a robust method for generating ESCs applicable to both permissive and nonpermissive mouse models, including the NOD mouse.
Main Methods:
- Utilized high concentrations of leukemia inhibitory factor (LIF) combined with serum replacement for ESC derivation.
- Applied the protocol to fresh and cryopreserved embryos from C57Bl/6, Balb/K, CBA/Ca, and NOD mice.
- Assessed ESC pluripotency, karyotype stability, in vivo teratoma formation, germline transmission in chimeras, and in vitro differentiation potential.
Main Results:
- Successfully derived novel ESC lines from C57Bl/6, Balb/K, CBA/Ca, and notably, the previously refractory NOD mice.
- NOD ESCs exhibited stable normal karyotypes, formed teratomas, produced germline-competent chimeras, and differentiated into embryoid bodies and dendritic cells.
- CBA/Ca ESCs demonstrated significant contribution to somatic germ layers and high germline transmission competence.
Conclusions:
- The developed protocol effectively enables the derivation of pluripotent ESCs from genetically diverse and previously nonpermissive mouse strains, including NOD mice.
- These novel ESC lines are valuable tools for modeling human diseases like type I diabetes and for generating specific cell types for research.
- The protocol's applicability to cryopreserved embryos enhances its utility for broad research applications.
