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An Efficient In Vitro Transposition Method by a Transcriptionally Regulated Sleeping Beauty System Packaged into an Integration Defective Lentiviral Vector
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Biologia
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JoVE Journal Biologia
An Efficient In Vitro Transposition Method by a Transcriptionally Regulated Sleeping Beauty System Packaged into an Integration Defective Lentiviral Vector

An Efficient In Vitro Transposition Method by a Transcriptionally Regulated Sleeping Beauty System Packaged into an Integration Defective Lentiviral Vector

DOI:
10.3791/56742-v

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10:13 min

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January 12, 2018

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, ,
1Centre for Regenerative Medicine, Department of Life Sciences,University of Modena and Reggio Emilia

Capítulos

  • 00:05Título
  • 00:51Transduction of Human Primary Keratinocytes
  • 04:31Cytofluorimetric Analysis of Transduced Primary Keratinocytes
  • 06:24Quantitative RT-PCR (qRT-PCR)
  • 07:24Results: Sleeping Beauty System Achieves Stable Integration of a GOI in the Human Genome
  • 08:40Conclusion

Summary

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This protocol describes a method to achieve stable integration of a gene of interest into the human genome by the transcriptionally regulated Sleeping Beauty system. Preparation of the integration of defective lentiviral vectors, the in vitro transduction of human cells, and the molecular assay on transduced cells are reported.

Tags

Keywords: Sleeping Beauty SystemIntegration Defective Lentiviral VectorGene TherapyPrimary CellsTranscriptionally RegulatedTransposition MethodIn VitroHyperactive SB100X TransposaseGene Of InterestTetracycline-regulatedPolybrene

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