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Novel codon-optimized mini-intronic plasmid for efficient, inexpensive, and xeno-free induction of pluripotency

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Item Type:Article
Title:Novel codon-optimized mini-intronic plasmid for efficient, inexpensive, and xeno-free induction of pluripotency
Creators Name:Diecke, S. and Lu, J. and Lee, J. and Termglinchan, V. and Kooreman, N.G. and Burridge, P.W. and Ebert, A.D. and Churko, J.M. and Sharma, A. and Kay, M.A. and Wu, J.C.
Abstract:The development of human induced pluripotent stem cell (iPSC) technology has revolutionized the regenerative medicine field. This technology provides a powerful tool for disease modeling and drug screening approaches. To circumvent the risk of random integration into the host genome caused by retroviruses, non-integrating reprogramming methods have been developed. However, these techniques are relatively inefficient or expensive. The mini-intronic plasmid (MIP) is an alternative, robust transgene expression vector for reprogramming. Here we developed a single plasmid reprogramming system which carries codon-optimized (Co) sequences of the canonical reprogramming factors (Oct4, Klf4, Sox2, and c-Myc) and short hairpin RNA against p53 ("4-in-1 CoMiP"). We have derived human and mouse iPSC lines from fibroblasts by performing a single transfection. Either independently or together with an additional vector encoding for LIN28, NANOG, and GFP, we were also able to reprogram blood-derived peripheral blood mononuclear cells (PBMCs) into iPSCs. Taken together, the CoMiP system offers a new highly efficient, integration-free, easy to use, and inexpensive methodology for reprogramming. Furthermore, the CoMIP construct is color-labeled, free of any antibiotic selection cassettes, and independent of the requirement for expression of the Epstein-Barr Virus nuclear antigen (EBNA), making it particularly beneficial for future applications in regenerative medicine.
Keywords:Cellular Reprogramming, Codon, Cultured Cells, Epstein-Barr Virus Nuclear Antigens, Fibroblasts, Fluorescence Microscopy, Homeodomain Proteins, Innate Immunity, Introns, Karyotyping, Mononuclear Leukocytes, Plasmids, Pluripotent Stem Cells, Proto-Oncogene Proteins c-myc, RNA-Binding Proteins, Small Interfering RNA, Transcription Factors, Tumor Suppressor Protein p53, Animals, Mice
Source:Scientific Reports
ISSN:2045-2322
Publisher:Nature Publishing Group
Volume:5
Page Range:8081
Date:28 January 2015
Official Publication:https://doi.org/10.1038/srep08081
PubMed:View item in PubMed

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