Rapid and Efficient Generation of Transgene-Free iPSC from a Small Volume of Cryopreserved Blood.
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Human peripheral blood and umbilical cord blood represent attractive sources of cells for reprogramming to induced pluripotent stem cells (iPSCs). However, to date, most of the blood-derived iPSCs were generated using either integrating methods or starting from T-lymphocytes that have genomic rearrangements thus bearing uncertain consequences when using iPSC-derived lineages for disease modeling and cell therapies. Recently, both peripheral blood and cord blood cells have been reprogrammed into transgene-free iPSC using the Sendai viral vector. Here we demonstrate that peripheral blood can be utilized for medium-throughput iPSC production without the need to maintain cell culture prior to reprogramming induction. Cell reprogramming can also be accomplished with as little as 3000 previously cryopreserved cord blood cells under feeder-free and chemically defined Xeno-free conditions that are compliant with standard Good Manufacturing Practice (GMP) regulations. The first iPSC colonies appear 2-3 weeks faster in comparison to previous reports. Notably, these peripheral blood- and cord blood-derived iPSCs are free of detectable immunoglobulin heavy chain (IGH) and T cell receptor (TCR) gene rearrangements, suggesting they did not originate from B- or T- lymphoid cells. The iPSCs are pluripotent as evaluated by the scorecard assay and in vitro multi lineage functional cell differentiation. Our data show that small volumes of cryopreserved peripheral blood or cord blood cells can be reprogrammed efficiently at a convenient, cost effective and scalable way. In summary, our method expands the reprogramming potential of limited or archived samples either stored at blood banks or obtained from pediatric populations that cannot easily provide large quantities of peripheral blood or a skin biopsy.
Pluripotent Stem Cells
Proto-Oncogene Proteins c-myc
Cell Culture Techniques
Octamer Transcription Factor-3
Kruppel-Like Transcription Factors
SOXB1 Transcription Factors
Induced Pluripotent Stem Cells
Kruppel-Like Factor 4
Published Version (Please cite this version)10.1007/s12015-015-9586-8
Publication InfoZhou, Hongyan; Martinez, Hector; Sun, Bruce; Li, Aiqun; Zimmer, Matthew; Katsanis, Nicholas; ... Chang, Stephen (2015). Rapid and Efficient Generation of Transgene-Free iPSC from a Small Volume of Cryopreserved Blood. Stem cell reviews and reports, 11(4). pp. 652-665. 10.1007/s12015-015-9586-8. Retrieved from https://hdl.handle.net/10161/24630.
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Erica Ellen Davis
Associate Professor of Pediatrics
Two key questions thematically underscore my research in the Center for Human Disease Modeling at Duke University: First of all, how can variation at the DNA level be functionally interpreted beyond the resolution of genetics arguments alone? Secondly, once empowered with functional information about genetic variants, how can pathogenic alleles be mapped back to disease phenotypes? Using the ciliary disease module as a model system of investigation, we are using multidisciplinary tactics to addr
Jean and George W. Brumley, Jr., M.D. Professor of Developmental Biology
Jerome S. Harris Distinguished Professor of Pediatrics
Dr. Kurtzberg is an internationally renowned expert in pediatric hematology/oncology, pediatric blood and marrow transplantation, umbilical cord blood banking and transplantation, and novel applications of cord blood and birthing tissues in the emerging fields of cellular therapies and regenerative medicine. Dr. Kurtzberg serves as the Director of the Marcus Center for Cellular Cures (MC3), Director of the Pediatric Transplant and Cellular Therapy Program, Director of the Carolina
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