Efficient Modulation of TP53 Expression in Human Induced Pluripotent Stem Cells.
Curr Protoc Stem Cell Biol
; 52(1): e102, 2020 03.
Article
en En
| MEDLINE
| ID: mdl-31883435
TP53 point mutations are found in 50% of all cancers and seem to play an important role in cancer pathogenesis. Thus, human induced pluripotent stem cells (hiPSCs) overexpressing mutant TP53 are a valuable tool for the generation of in vitro models of cancer stem cells or for in vivo xenograft models. Here, we describe a protocol for the alteration of gene expression in hiPSCs via overexpression of a mutant form of the TP53 (R249S) gene using lentiviral transduction. A high amount of TP53 protein is detected 1 week after transduction and antibiotic selection. Differentiation of transduced hiPSCs gives insight into better understanding cancer formation in different tissues and may be a useful tool for genetic or pharmacologic screening assays. © 2019 The Authors. Basic Protocol 1: Production and concentration of third-generation lentivirus Support Protocol 1: Cloning of gene of interest into modulation vector Support Protocol 2: Preparation of DMEM GlutaMAX™ with 10% fetal bovine serum and 1% penicillin-streptomycin Basic Protocol 2: Transduction of human induced pluripotent stem cells and selection of positively transfected cells Support Protocol 3: Preparation of Matrigel® -coated plates Support Protocol 4: Preparation of mTeSR™1 medium.
Palabras clave
Texto completo:
1
Bases de datos:
MEDLINE
Asunto principal:
Expresión Génica
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Proteína p53 Supresora de Tumor
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Técnicas de Cultivo de Célula
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Células Madre Pluripotentes Inducidas
Límite:
Humans
Idioma:
En
Revista:
Curr Protoc Stem Cell Biol
Año:
2020
Tipo del documento:
Article
País de afiliación:
Alemania