title: EOS lentiviral vector selection system for human induced pluripotent stem cells.
abstract: Generation of induced pluripotent stem (iPS) cells from patients has exciting applications for studying molecular mechanisms of diseases, screening drugs and ultimately for use in cell therapies. However, the low efficiency and heterogeneous nature of reprogramming is a major impediment to the generation of personalized iPS cell lines. We reported in Nature Methods (6, 370-376, 2009) the first selection system to enrich for reprogrammed human iPS cells. Using a lentiviral vector that specifically expresses the enhanced green fluorescence protein and puromycin resistance genes in pluripotent stem cells, it is now possible to mark and enrich for human iPS cell colonies expressing endogenous pluripotency markers. In this study, we describe a detailed protocol for the production of the pluripotent state-specific lentiviral vector and the selection system for the induction of healthy and disease-specific human iPS cells. Overall, preparation of the selection system takes 2 weeks, and the generation of human iPS cells takes approximately 2 months.
authors.affiliation: Developmental and Stem Cell Biology Program, SickKids, Toronto, Ontario, Canada.
authors.forename: Akitsu
authors.initials: A
authors.lastname: Hotta
authors.forename: Aaron Y L
authors.initials: AY
authors.lastname: Cheung
authors.forename: Natalie
authors.initials: N
authors.lastname: Farra
authors.forename: Kamal
authors.initials: K
authors.lastname: Garcha
authors.forename: Wing Y
authors.initials: WY
authors.lastname: Chang
authors.forename: Peter
authors.initials: P
authors.lastname: Pasceri
authors.forename: William L
authors.initials: WL
authors.lastname: Stanford
authors.forename: James
authors.initials: J
authors.lastname: Ellis
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D018929:Cell Culture Techniques; D004351:Drug Resistance / Q000235:genetics; D005347:Fibroblasts / Q000166:cytology / Q000821:virology; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000378:metabolism; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000166:cytology* / Q000821:virology; D016086:Lentivirus / Q000235:genetics*; D051379:Mice; D014162:Transfection / Q000379:methods
chemical_list: D015415:Biomarkers; D049452:Green Fluorescent Proteins