title: Simple derivation of transgene-free iPS cells by a dual recombinase approach.
abstract: Mammalian cells can be reprogrammed into induced pluripotent stem cells (iPSCs), a valuable tool for in vitro disease modeling and regenerative medicine. These applications demand for iPSCs devoid of reprogramming factor transgenes, but current procedures for the derivation of transgene-free iPSCs are inefficient and cumbersome. Here, we describe a new approach for the simple derivation of transgene-free iPSCs by the sequential use of two DNA recombinases, C31 Integrase and Cre, to control the genomic insertion and excision of a single, non-viral reprogramming vector. We show that such transgene-free iPSCs exhibit gene expression profiles and pluripotent developmental potential comparable to genuine, blastocyst-derived embryonic stem cells. As shown by a reporter iPSC line for the differentiation into midbrain dopaminergic neurons, the dual recombinase approach offers a simple and efficient way to derive transgene-free iPSCs for studying disease mechanisms and cell replacement therapies.
authors.affiliation: Institute of Developmental Genetics, Helmholtz Zentrum München, 85764, Munich, Germany, anna.pertek@helmholtz-muenchen.de.
authors.forename: Anna
authors.initials: A
authors.lastname: Pertek
authors.forename: Florian
authors.initials: F
authors.lastname: Meier
authors.forename: Martin
authors.initials: M
authors.lastname: Irmler
authors.forename: Johannes
authors.initials: J
authors.lastname: Beckers
authors.forename: Stavroula
authors.initials: S
authors.lastname: Skylaki
authors.forename: Max
authors.initials: M
authors.lastname: Endele
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Wurst
authors.forename: Nilima
authors.initials: N
authors.lastname: Prakash
authors.forename: Ralf
authors.initials: R
authors.lastname: Kühn
mesh_terms: D000818:Animals; D001709:Biotechnology; D002454:Cell Differentiation; D002478:Cells, Cultured; D065150:Cellular Reprogramming / Q000235:genetics; D059290:Dopaminergic Neurons / Q000166:cytology / Q000378:metabolism; D005260:Female; D017930:Genes, Reporter; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics; D057026:Induced Pluripotent Stem Cells / Q000166:cytology* / Q000378:metabolism* / Q000637:transplantation; D019426:Integrases / Q000235:genetics; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008822:Mice, Transgenic; D059467:Transcriptome
chemical_list: D049452:Green Fluorescent Proteins; C045073:Cre recombinase; D019426:Integrases