Stable generation of serum- and feeder-free embryonic stem cell-derived mice with full germline-competency by using a GSK3 specific inhibitor.
C57BL/6 (B6)-derived embryonic stem (ES) cells are not widely used to generate knockout mice despite the advantage of a well-defined genetic background because of poor developmental potential. We newly established serum- and feeder-free B6 ES cells with full developmental potential by using leukemia inhibitory factor (LIF) and 6-bromoindirubin-3'-oxime (BIO), a glycogen synthase kinase-3 (GSK3) inhibitor. BIO treatment significantly increased the expression levels of 364 genes including pluripotency markers such as Nanog and Klf family. Unexpectedly, by aggregating or microinjecting those ES cells to each eight-cell-stage diploid embryo, we stably generated germline-competent ES-derived mice. Furthermore, founder mice completely derived from female XO, heterozygous, or homozygous mutant B6 ES cells were directly available for intercross breeding and phenotypic analysis. We hereby propose that serum- and feeder-free B6 ES cells stimulated with LIF plus GSK3 inhibitor are valuable for generating mouse models on B6 background.
Sato
Hiromu
H
Tsukuba Research Institute, Banyu Pharmaceutical Co., Ltd., Tsukuba, Japan.
Amagai
Keiko
K
Shimizukawa
Rie
R
Tamai
Yoshitaka
Y
D000818:Animals; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D004622:Embryo, Mammalian / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D053595:Embryonic Stem Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D005260:Female; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors / Q000378:metabolism; D007211:Indoles / Q000494:pharmacology*; D053605:Leukemia Inhibitory Factor / Q000494:pharmacology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008813:Mice, Inbred ICR; D008815:Mice, Inbred Strains; D018345:Mice, Knockout; D020411:Oligonucleotide Array Sequence Analysis; D010091:Oximes / Q000494:pharmacology*; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D054411:Receptors, Leptin / Q000235:genetics / Q000378:metabolism; D039901:Totipotent Stem Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism
C483321:6-bromoindirubin-3'-oxime; D016895:Culture Media, Serum-Free; D007211:Indoles; D053605:Leukemia Inhibitory Factor; D010091:Oximes; D054411:Receptors, Leptin; D038362:Glycogen Synthase Kinase 3