Isolation of human iPS cells using EOS lentiviral vectors to select for pluripotency.
Induced pluripotent stem (iPS) cells may be of use in regenerative medicine. However, the low efficiency of reprogramming is a major impediment to the generation of patient-specific iPS cell lines. Here we report the first selection system for the isolation of human iPS cells. We developed the EOS (Early Transposon promoter and Oct-4 (Pou5f1) and Sox2 enhancers) lentiviral vector to specifically express in mouse and human embryonic stem cells but not in primary fibroblasts. The bicistronic EOS vector marked emerging mouse and human iPS cell colonies with EGFP, and we used puromycin selection to aid the isolation of iPS cell lines that expressed endogenous pluripotency markers. These lines differentiated into cell types from all three germ layers. Reporter expression was extinguished upon differentiation and therefore monitored the residual pluripotent cells that form teratomas. Finally, we used EOS selection to establish Rett syndrome-specific mouse and human iPS cell lines with known mutations in MECP2.
Hotta
Akitsu
A
Developmental and Stem Cell Biology Program, Toronto, Ontario, Canada.
Cheung
Aaron Y L
AY
Farra
Natalie
N
Vijayaragavan
Kausalia
K
Séguin
Cheryle A
CA
Draper
Jonathan S
JS
Pasceri
Peter
P
Maksakova
Irina A
IA
Mager
Dixie L
DL
Rossant
Janet
J
Bhatia
Mickie
M
Ellis
James
J
D000818:Animals; D054337:Cell Dedifferentiation / Q000235:genetics*; D002454:Cell Differentiation / Q000187:drug effects; D002469:Cell Separation / Q000379:methods*; D004251:DNA Transposable Elements / Q000235:genetics; D004622:Embryo, Mammalian / Q000166:cytology; D053595:Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism; D004742:Enhancer Elements, Genetic / Q000235:genetics; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D017930:Genes, Reporter / Q000235:genetics*; D005822:Genetic Vectors / Q000235:genetics*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D016086:Lentivirus / Q000235:genetics*; D051783:Methyl-CpG-Binding Protein 2 / Q000235:genetics; D051379:Mice; D016688:Mice, Inbred NOD; D008817:Mice, Mutant Strains; D016513:Mice, SCID; D039904:Pluripotent Stem Cells / Q000166:cytology* / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D015518:Rett Syndrome / Q000235:genetics / Q000473:pathology; D013724:Teratoma / Q000473:pathology
D004251:DNA Transposable Elements; D051783:Methyl-CpG-Binding Protein 2; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins