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Frog Prince transposon-based RNAi vectors mediate efficient gene knockdown in human cells

Authors

  • C.D. Kaufman
  • Z. Izsvak
  • A. Katzer
  • Z. Ivics

Journal

  • Journal of RNAi and Gene Silencing

Citation

  • J RNAi Gene Silencing 1 (2): 97-104

Abstract

  • We have developed a stable RNA interference (RNAi) delivery system that is based on the Frog Prince transposable element. This plasmid-based vector system combines the gene silencing capabilities of H1 polymerase III promoter-driven short hairpin RNAs (shRNA) with the advantages of stable and efficient genomic integration of the shRNA cassette mediated by transposition. We show that the Frog Prince-based shRNA expressing system can efficiently knock down the expression of both exogenous as well as endogenous genes in human cells. Furthermore, we use the Frog Prince-based system to study the effect of knockdown of the DNA repair factor Ku70 on transposition of the Sleeping Beauty transposon. Transposon-mediated genomic integration ensures that the shRNA expression cassette and a selectable marker gene within the transposon remain intact and physically linked. We demonstrate that a major advantage of our vector system over plasmid-based shRNA delivery is both its enhanced frequency of intact genomic integration as well as higher target suppression in transgenic human cells. Due to its simplicity and effectiveness, transposon-based RNAi is an emerging tool to facilitate analysis of gene function through the establishment of stable loss-of-function cell lines.


DOI

doi:RNAiJ-Issues/Issue-2/AuthorsIndex-2.pdf