title: Gene Transfer into Pluripotent Stem Cells via Lentiviral Transduction.
abstract: Recombinant lentiviral vectors are powerful tools to stably manipulate human pluripotent stem cells. They can be used to deliver ectopic genes, shRNAs, miRNAs, or any possible genetic DNA sequence into diving and nondividing cells. Here we describe a general protocol for the production of self-inactivating lentiviral vector particles and their purification to high titers by either ultracentrifugation or ultrafiltration. Next we provide a basic procedure to transduce human pluripotent stem cells and propagate clonal cell lines.
authors.affiliation: Institute of Clinical Biochemistry, Hannover Medical School, 30625, Hannover, Germany. naujok.ortwin@mh-hannover.de.
authors.forename: Ortwin
authors.initials: O
authors.lastname: Naujok
authors.affiliation: Institute of Clinical Biochemistry, Hannover Medical School, 30625, Hannover, Germany.
authors.forename: Ulf
authors.initials: U
authors.lastname: Diekmann
authors.affiliation: Institute of Clinical Biochemistry, Hannover Medical School, 30625, Hannover, Germany.
authors.forename: Matthias
authors.initials: M
authors.lastname: Elsner
mesh_terms: D018929:Cell Culture Techniques / Q000379:methods; D002478:Cells, Cultured; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000235:genetics* / Q000302:isolation & purification; D057809:HEK293 Cells; D006801:Humans; D016086:Lentivirus / Q000235:genetics* / Q000302:isolation & purification; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000378:metabolism*; D014161:Transduction, Genetic / Q000379:methods*; D014461:Ultracentrifugation / Q000379:methods; D014462:Ultrafiltration / Q000379:methods
keywords: Human pluripotent stem cells; Lentivirus production; Primed pluripotent stem cells; Selection of cell clones; Transduction