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34 - Generation of highly specific vector-based shRNAi libraries directed against the entire human genome
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- By Makoto Miyagishi, Department of Chemistry and Biotechnology, School of Engineering, The University of Tokyo, Sahohime Matsumoto, Department of Chemistry and Biotechnology, School of Engineering, The University of Tokyo, Takashi Futami, Department of Chemistry and Biotechnology, School of Engineering, The University of Tokyo, Hideo Akashi, Department of Chemistry and Biotechnology, School of Engineering, The University of Tokyo, Krishnarao Appasani, GeneExpression Systems, Inc., Yasuomi Takagi, iGENE Therapeutics, Inc., Shizuyo Sutou, iGENE Therapeutics, Inc., Takashi Kadowaki, Department of Internal Medicine, Graduate School of Medicine, The University of Tokyo, Ryozo Nagai, Department of Internal Medicine, Graduate School of Medicine, The University of Tokyo, Kazunari Taira, Department of Chemistry and Biotechnology, School of Engineering, The University of Tokyo
- Edited by Krishnarao Appasani, GeneExpression Systems, Inc., Massachusetts
- Foreword by Andrew Fire, Stanford University, California, Marshall Nirenberg
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- Book:
- RNA Interference Technology
- Published online:
- 31 July 2009
- Print publication:
- 17 January 2005, pp 480-496
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- Chapter
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Summary
Introduction
The success of the Human Genome Project and the availability of the complete sequence of the human genome have revealed the existence of numerous genes whose functions are unknown. Although many functional genes were successfully identified using libraries of randomized ribozymes (Kruger et al., 2000; Li et al., 2000; Welch et al., 2000; Beger et al., 2001; Kawasaki et al., 2002; Kawasaki and Taira, 2002; Onuki et al., 2002; Nelson et al., 2003; Rhoades and Wong-Staal, 2003; Suyama et al., 2003a, b; Chatterton et al., 2004; Kuwabara et al., 2004; Onuki et al., 2004), the randomized library naturally contained many ribozymes that could not hybridize with any human gene transcripts, resulting in some false positives. While exploitation of RNA interference (RNAi) is hampered by the induction of the interferon response upon the introduction of double-stranded RNA (dsRNA) into mammalian cells (Elbashir et al., 2001), RNAi can be a powerful tool in gene analysis and, for example, the functions of a number of genes in the nematode Caenorhabditis elegans have been identified as being associated with particular mutant phenotypes (Fraser et al., 2000; Gonczy et al., 2000). Recent progress in mammalian cells, as discussed in this chapter, suggests that it should soon be possible to extend genome-wide approaches to mammalian cells, using libraries of small interfering RNA (siRNA) oligonucleotides or siRNA-expression libraries.
Efforts to exploit the phenomenon of RNA interference (RNAi) in mammalian cells have been hampered by the interferon response in cells into which foreign RNA is introduced.