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Multiple shRNA expressing vector enhances efficiency of gene silencing

  • Song, Jun (Division of Oral Biology and Medicine, UCLA School of Dentistry) ;
  • Giang, An (Division of Oral Biology and Medicine, UCLA School of Dentistry) ;
  • Lu, Yingchun (Division of Oral Biology and Medicine, UCLA School of Dentistry) ;
  • Pang, Shen (Division of Oral Biology and Medicine, UCLA School of Dentistry) ;
  • Chiu, Robert (Division of Oral Biology and Medicine, UCLA School of Dentistry)
  • Published : 2008.05.31

Abstract

RNA interference (RNAi) is the process of sequence-specific gene silencing. However, RNAi efficiency still needs to be improved for effective inhibition of target genes. We have developed an effective strategy to express multiple shRNAs (small hairpin RNA) simultaneously using multiple RNA Polymerase III (Pol III) promoters in a single vector. Our data demonstrate that multiple shRNAs expressed from Pol III promoters have a synergistic effect in repressing the target gene. Silencing of endogenous cyclophilin A (CypA) or key HIV viral genes by multiple shRNAs results in significant inhibition of the target gene.

Keywords

References

  1. Shi, Y. (2003) Mammalian RNAi for the masses. Trends Genet. 19, 9-12 https://doi.org/10.1016/S0168-9525(02)00005-7
  2. Reynolds, A., Leake, D., Boese, Q., Scaringe, S., Marshall, W. S. and Khvorova, A. (2004) Rational siRNA design for RNA interference. Nat. Biotechnol. 22, 326-330 https://doi.org/10.1038/nbt936
  3. Gonzalez, S., Castanotto, D., Li, H., Olivares, S., Jensen, M. C., Forman, S. J., Rossi, J. J. and Cooper, L. J. (2005) Amplification of RNAi-targeting HLA mRNAs. Mol. Ther. 11, 811-818 https://doi.org/10.1016/j.ymthe.2004.12.023
  4. Song, J., Lu, Y. C., Yokoyama, K., Rossi, J. J. and Chiu, R. (2004) Cyclophilin A is required for retinoic acid-induced neuronal differentiation in p19 cells. J. Biol. Chem. 279, 24414-24419 https://doi.org/10.1074/jbc.M311406200
  5. Stram, Y. and Kuzntzova, L. (2006) Inhibition of viruses by RNA interference. Virus Genes. 32, 299-306 https://doi.org/10.1007/s11262-005-6914-0
  6. van Rij, R.P. and Andino, R. (2006) The silent treatment: RNAi as a defense against virus infection in mammals. Trends Biotechnol. 24, 186-193 https://doi.org/10.1016/j.tibtech.2006.02.006
  7. Sun, D., Melegari, M., Sridhar, S., Rogler, C. E. and Zhu, L. (2006) Multi-miRNA hairpin method that improves gene knockdown efficiency and provides linked multigene knockdown. BioTechniques 41, 59-63 https://doi.org/10.2144/000112203
  8. Xia, X., Zhou, H. and Xu, Z. (2006) Multiple shRNAs expressed by an inducible pol II promoter can knock down the expression of multiple target genes. BioTechniques 41, 64-68 https://doi.org/10.2144/000112198
  9. Marks, A. R. (1996) Cellular functions of immunophilins. Physiol. Rev. 76, 631-649 https://doi.org/10.1152/physrev.1996.76.3.631

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