RNA interference is responsible for reduction of transgene expression after Sleeping Beauty transposase mediated somatic integration.

PloS One
Christina Rauschhuber, Anja Ehrhardt

Abstract

Integrating non-viral vectors based on transposable elements are widely used for genetically engineering mammalian cells in functional genomics and therapeutic gene transfer. For the Sleeping Beauty (SB) transposase system it was demonstrated that convergent transcription driven by the SB transposase inverted repeats (IRs) in eukaryotic cells occurs after somatic integration. This could lead to formation of double-stranded RNAs potentially presenting targets for the RNA interference (RNAi) machinery and subsequently resulting into silencing of the transgene. Therefore, we aimed at investigating transgene expression upon transposition under RNA interference knockdown conditions. To establish RNAi knockdown cell lines we took advantage of the P19 protein, which is derived from the tomato bushy stunt virus. P19 binds and inhibits 21 nucleotides long, small-interfering RNAs and was shown to sufficiently suppress RNAi. We found that transgene expression upon SB mediated transposition was enhanced, resulting into a 3.2-fold increased amount of colony forming units (CFU) after transposition. In contrast, if the transgene cassette is insulated from the influence of chromosomal position effects by the chicken-derived cHS4 insulating seq...Continue Reading

References

Dec 20, 1991·Journal of Molecular Biology·S Kuhstoss, R N Rao
Nov 26, 1999·Proceedings of the National Academy of Sciences of the United States of America·O VoinnetD C Baulcombe
May 10, 2000·Proceedings of the National Academy of Sciences of the United States of America·A C GrothM P Calos
Nov 15, 2000·Proceedings of the National Academy of Sciences of the United States of America·C LlaveJ C Carrington
Feb 7, 2001·Genes & Development·S M ElbashirT Tuschl
Mar 27, 2002·Genes & Development·Zissimos MourelatosGideon Dreyfuss
Oct 16, 2002·Nature Biotechnology·Eric C OlivaresMichele P Calos
Aug 9, 2003·Current Biology : CB·Nadine L VastenhouwRonald H A Plasterk
Dec 19, 2003·Genes & Development·Rui YiBryan R Cullen
May 1, 2004·Science·Sébastien PfefferThomas Tuschl
Jun 29, 2004·Trends in Genetics : TIG·Nadine L Vastenhouw, Ronald H A Plasterk
Jul 21, 2004·Molecular Cell·Gunter MeisterThomas Tuschl
Aug 7, 2004·Science·Kevin V MorrisDavid J Looney
Sep 10, 2004·Nucleic Acids Research·Yan Zeng, Bryan R Cullen
Jul 15, 2005·Journal of Virology·M Gunnar AnderssonGöran Akusjärvi
Aug 25, 2005·Cold Spring Harbor Symposia on Quantitative Biology·V J P RobertR H A Plasterk
Oct 21, 2005·BioTechniques·Rui Shi, Vincent L Chiang
Mar 7, 2006·Nature Reviews. Microbiology·Herman B Scholthof
Aug 29, 2006·Nature Structural & Molecular Biology·Daniel H KimJohn J Rossi
Aug 29, 2006·Nature Structural & Molecular Biology·Nuo Yang, Haig H Kazazian
Aug 29, 2006·Nature Structural & Molecular Biology·Bethany A JanowskiDavid R Corey
Dec 8, 2006·Nucleic Acids Symposium Series·Hiroaki Kawasaki, Kazunari Taira
Dec 14, 2006·Molecular Therapy : the Journal of the American Society of Gene Therapy·Anja EhrhardtMark A Kay
Mar 9, 2007·Nucleic Acids Research·Stephen R YantMark A Kay
Oct 17, 2007·Molecular and Cellular Biology·Brian S GarrisonMark A Kay
Dec 12, 2007·Molecular Therapy : the Journal of the American Society of Gene Therapy·Oliver WaliskoZoltán Ivics
Feb 4, 2009·Cell Research·Ingela Djupedal, Karl Ekwall

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Citations

Dec 11, 2013·Journal of Biomedical Science·Kristian Alsbjerg SkipperJacob Giehm Mikkelsen
Jan 15, 2013·Translational Research : the Journal of Laboratory and Clinical Medicine·Perry B HackettLaurence J N Cooper
Oct 5, 2016·Critical Reviews in Biochemistry and Molecular Biology·Suneel A NarayanavariZsuzsanna Izsvák
Jun 25, 2015·Microbiology Spectrum·Zoltán Ivics, Zsuzsanna Izsvák

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Methods Mentioned

BETA
PCR
transfection
fluorescence-activated cell sorting
flow cytometry
immunoprecipitation
fluorescent
activated cell sorting
FACS
Reverse Transcription PCR

Software Mentioned

FACS Diva

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