Stable production of mutant mice from double gene converted ES cells with puromycin and neomycin.
The antibiotic puromycin is an effective inhibitor of protein synthesis and puromycin N-acetyl transferase gene could be used as a dominant selection marker. We report the effective production of mutant mice from double gene-converted ES cells by selection with G418 and puromycin. We confirmed that (i) puromycin efficiently inhibited the growth of ES cells at a low-dose (0.1 microgram/ml) and for a short time (2 days), independent of G418 selection; (ii) when these selected ES cells were injected into eight-cell stage embryos, the cells produced chimeras with high levels of chimerism; (iii) these chimeric males were fertile and exclusively yielded ES cell-derived offspring; and (iv) each offspring contained both neomycin transferase and puromycin N-acetyl transferase genes.
Watanabe
S
S
Laboratory of Neurobiology and Behavioral Genetics, National Institute for Physiological Sciences, Okazaki, Japan.
Kai
N
N
Yasuda
M
M
Kohmura
N
N
Sanbo
M
M
Mishina
M
M
Yagi
T
T
D000123:Acetyltransferases / Q000096:biosynthesis / Q000235:genetics*; D000199:Actins / Q000235:genetics; D000818:Animals; D001483:Base Sequence; D002455:Cell Division / Q000187:drug effects; D002645:Chickens; D002678:Chimera; D017931:DNA Primers; D004622:Embryo, Mammalian; D004625:Embryo, Nonmammalian; D005260:Female; D016259:Genes, myc; D005839:Gentamicins / Q000494:pharmacology; D006801:Humans; D008297:Male; D051379:Mice; D008815:Mice, Inbred Strains; D008817:Mice, Mutant Strains; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D009355:Neomycin / Q000494:pharmacology*; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins / Q000235:genetics; D051076:Proto-Oncogene Proteins c-fyn; D011519:Proto-Oncogenes; D011691:Puromycin / Q000494:pharmacology*; D013234:Stem Cells / Q000166:cytology / Q000187:drug effects / Q000502:physiology*
D000199:Actins; D017931:DNA Primers; D005839:Gentamicins; D011518:Proto-Oncogene Proteins; D011691:Puromycin; C010680:antibiotic G 418; D000123:Acetyltransferases; C045336:puromycin N-acetyltransferase; C494957:FYN protein, human; C494960:Fyn protein, mouse; D051076:Proto-Oncogene Proteins c-fyn; D009355:Neomycin