Please use this identifier to cite or link to this item: https://hdl.handle.net/10356/103690
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dc.contributor.authorMakhija, Harshyaaen
dc.contributor.authorRoy, Sukien
dc.contributor.authorHoon, Shawnen
dc.contributor.authorGhadessy, Farid Johnen
dc.contributor.authorWong, Desmonden
dc.contributor.authorJaiswal, Rahulen
dc.contributor.authorCampana, Darioen
dc.contributor.authorDröge, Peteren
dc.date.accessioned2019-01-04T07:50:17Zen
dc.date.accessioned2019-12-06T21:18:03Z-
dc.date.available2019-01-04T07:50:17Zen
dc.date.available2019-12-06T21:18:03Z-
dc.date.issued2018en
dc.identifier.citationMakhija, H., Roy, S., Hoon, S., Ghadessy, F. J., Wong, D., Jaiswal, R., . . . Dröge, P. (2018). A novel λ integrase-mediated seamless vector transgenesis platform for therapeutic protein expression. Nucleic Acids Research, 46(16), e99-. doi:10.1093/nar/gky500en
dc.identifier.issn0305-1048en
dc.identifier.urihttps://hdl.handle.net/10356/103690-
dc.description.abstractAdvances in stem cell engineering, gene therapy and molecular medicine often involve genome engineering at a cellular level. However, functionally large or multi transgene cassette insertion into the human genome still remains a challenge. Current practices such as random transgene integration or targeted endonuclease-based genome editing are suboptimal and might pose safety concerns. Taking this into consideration, we previously developed a transgenesis tool derived from phage λ integrase (Int) that precisely recombines large plasmid DNA into an endogenous sequence found in human Long INterspersed Elements-1 (LINE-1). Despite this advancement, biosafety concerns associated with bacterial components of plasmids, enhanced uptake and efficient transgene expression remained problematic. We therefore further improved and herein report a more superior Int-based transgenesis tool. This novel Int platform allows efficient and easy derivation of sufficient amounts of seamless supercoiled transgene vectors from conventional plasmids via intramolecular recombination as well as subsequent intermolecular site-specific genome integration into LINE-1. Furthermore, we identified certain LINE-1 as preferred insertion sites for Int-mediated seamless vector transgenesis, and showed that targeted anti-CD19 chimeric antigen receptor gene integration achieves high-level sustained transgene expression in human embryonic stem cell clones for potential downstream therapeutic applications.en
dc.description.sponsorshipASTAR (Agency for Sci., Tech. and Research, S’pore)en
dc.description.sponsorshipNMRC (Natl Medical Research Council, S’pore)en
dc.format.extent14 p.en
dc.language.isoenen
dc.relation.ispartofseriesNucleic Acids Researchen
dc.rights© 2018 The Author(s). Published by Oxford University Press on behalf of Nucleic Acids Research. This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.comen
dc.subjectTransgenesisen
dc.subjectProtein Expressionen
dc.subjectDRNTU::Science::Biological sciencesen
dc.titleA novel λ integrase-mediated seamless vector transgenesis platform for therapeutic protein expressionen
dc.typeJournal Articleen
dc.contributor.schoolSchool of Biological Sciencesen
dc.contributor.researchNTU Institute of Structural Biologyen
dc.identifier.doi10.1093/nar/gky500en
dc.description.versionPublished versionen
item.fulltextWith Fulltext-
item.grantfulltextopen-
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