Design and synthesis of small interfering RNA (siRNA)

doi:10.1017/CBO9780511546402.009

6 - Design and synthesis of small interfering RNA (siRNA)

Published online by Cambridge University Press: 31 July 2009

Queta Boese ,

William S. Marshall and

Anastasia Khvorova

Edited by

Krishnarao Appasani

Foreword by

Andrew Fire and

Marshall Nirenberg

Show author details

Queta Boese: Affiliation:
Dharmacon, Inc.
William S. Marshall: Affiliation:
Dharmacon, Inc.
Anastasia Khvorova: Affiliation:
Dharmacon, Inc.
Krishnarao Appasani: Affiliation:
GeneExpression Systems, Inc., Massachusetts
Andrew Fire: Affiliation:
Stanford University, California

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Summary

Efficient RNA interference (RNAi) depends on siRNA design and synthesis

RNAi is a powerful technology with tremendous utility for functional genomic analysis, drug discovery strategies and therapeutic applications (Appasani 2003). While this pathway for post-transcriptional gene regulation is ubiquitous among eukaryotes, species-specific variations in the mechanism impact the utility of this pathway. These species-specific distinctions have strong implications with regard to the design, production, and delivery of the functional silencing intermediates. For example, in Caenorhabditis elegans (C. elegans), simple exposure by soaking (Tabara et al. 1998; Timmons and Fire 1998), feeding (Fraser et al. 2000; Timmons et al. 2001), or injecting (Fire et al. 1998) the nematode with long dsRNA is sufficient to induce prolonged and potent gene knockdown. Silencing efficiency appears to be due to siRNA-primed amplification of additional dsRNA from the mRNA target resulting in a secondary pool of Dicer processed duplexes (Sijen et al. 2001; Tijsterman et al. 2002). This mechanism is characteristic of post-transcriptional gene silencing in nematodes and other lower eukaryotes and is mediated by an RNA-dependent RNA polymerase [(RdRP) (Sijen et al. 2001; Martens et al. 2002)]. Invariably, several of the newly generated siRNAs will be capable of proficient gene-specific knockdown thereby eliminating the need to carefully design and synthesize a single siRNA silencing intermediate.

In mammalian cell culture models, preliminary attempts to induce RNAi using long dsRNA met with limited success (Tuschl et al. 1999; Caplen et al. 2000; Zhao et al. 2001).

Type: Chapter
Information: RNA Interference Technology
From Basic Science to Drug Development
, pp. 103 - 117

DOI: https://doi.org/10.1017/CBO9780511546402.009 [Opens in a new window]

Publisher: Cambridge University Press

Print publication year: 2005

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