Abstract
C57BL/6 mouse is the most standard strain in mouse genetics. The strain does, however, have several disadvantages; one being the difficulty in establishing embryonic stem (ES) cells. No reliable C57BL/6 ES cell line is widely available for creating mutant mice through gene targeting. It also greatly favors mouse genetics if one can routinely make multiple mutations by stably culturing germline-competent C57BL/6 ES cells or if one can routinely establish ES cells from C57BL/6-derived mutant mice to make multiple mutations. Recently, an ES culture method with three inhibitors (3i: SU5402 for FGFR, PD184352 for ERK, and CHIR99021 for GSK3) has been reported. Here we show that this 3i method is extremely instrumental in establishing and culturing germline-competent ES cells in the C57BL/6N strain.
MeSH terms
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Animals
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Benzamides / pharmacology
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Cell Culture Techniques / methods*
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Cell Differentiation / drug effects
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Cells, Cultured
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Embryo, Mammalian / cytology*
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Embryonic Stem Cells / cytology*
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Embryonic Stem Cells / drug effects
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Embryonic Stem Cells / metabolism
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Extracellular Signal-Regulated MAP Kinases / antagonists & inhibitors
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Female
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Fibroblasts / cytology*
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Fibroblasts / drug effects
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Fibroblasts / metabolism
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Gene Expression Profiling
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Germ Cells / cytology
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Germ Cells / drug effects
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Germ Cells / metabolism
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Glycogen Synthase Kinase 3 / antagonists & inhibitors
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Immunohistochemistry
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Karyotyping
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Male
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Mice
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Mice, Inbred C57BL
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Mice, Inbred Strains
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Pyridines / pharmacology
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Pyrimidines / pharmacology
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Pyrroles / pharmacology
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Receptors, Fibroblast Growth Factor / antagonists & inhibitors
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Recombination, Genetic
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Reverse Transcriptase Polymerase Chain Reaction
Substances
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2-(2-chloro-4-iodophenylamino)-N-cyclopropylmethoxy-3,4-difluorobenzamide
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Benzamides
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Chir 99021
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Pyridines
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Pyrimidines
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Pyrroles
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Receptors, Fibroblast Growth Factor
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SU 5402
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Extracellular Signal-Regulated MAP Kinases
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Glycogen Synthase Kinase 3