Integration-Free iPS Cells Engineered Using Human Artificial Chromosome Vectors
Open Access
- 5 October 2011
- journal article
- research article
- Published by Public Library of Science (PLoS) in PLOS ONE
- Vol. 6 (10), e25961
- https://doi.org/10.1371/journal.pone.0025961
Abstract
Human artificial chromosomes (HACs) have unique characteristics as gene-delivery vectors, including episomal transmission and transfer of multiple, large transgenes. Here, we demonstrate the advantages of HAC vectors for reprogramming mouse embryonic fibroblasts (MEFs) into induced pluripotent stem (iPS) cells. Two HAC vectors (iHAC1 and iHAC2) were constructed. Both carried four reprogramming factors, and iHAC2 also encoded a p53-knockdown cassette. iHAC1 partially reprogrammed MEFs, and iHAC2 efficiently reprogrammed MEFs. Global gene expression patterns showed that the iHACs, unlike other vectors, generated relatively uniform iPS cells. Under non-selecting conditions, we established iHAC-free iPS cells by isolating cells that spontaneously lost iHAC2. Analyses of pluripotent markers, teratomas and chimeras confirmed that these iHAC-free iPS cells were pluripotent. Moreover, iHAC-free iPS cells with a re-introduced HAC encoding Herpes Simplex virus thymidine kinase were eliminated by ganciclovir treatment, indicating that the HAC safeguard system functioned in iPS cells. Thus, the HAC vector could generate uniform, integration-free iPS cells with a built-in safeguard system.Keywords
This publication has 39 references indexed in Scilit:
- Induced Pluripotent Stem Cells Generated Without Viral IntegrationScience, 2008
- Correction of a genetic defect in multipotent germline stem cells using a human artificial chromosomeGene Therapy, 2008
- Pancreatic endoderm derived from human embryonic stem cells generates glucose-responsive insulin-secreting cells in vivoNature Biotechnology, 2008
- Sequential Expression of Pluripotency Markers during Direct Reprogramming of Mouse Somatic CellsCell Stem Cell, 2008
- Induced Pluripotent Stem Cell Lines Derived from Human Somatic CellsScience, 2007
- Induction of Pluripotent Stem Cells from Adult Human Fibroblasts by Defined FactorsCell, 2007
- Induction of Pluripotent Stem Cells from Mouse Embryonic and Adult Fibroblast Cultures by Defined FactorsCell, 2006
- Ectopic Expression of Oct-4 Blocks Progenitor-Cell Differentiation and Causes Dysplasia in Epithelial TissuesCell, 2005
- p53 induces differentiation of mouse embryonic stem cells by suppressing Nanog expressionNature Cell Biology, 2004
- Functional expression and germline atransmission of a human chromosome fragment in chimaeric miceNature Genetics, 1997