Mechanistic Insights Aid Computational Short Interfering RNA Design

Boese, Queta; Leake, Devin; Reynolds, Angela; Read, Steven; Scaringe, Stephen A.; Marshall, William S.; Khvorova, Anastasia

doi:10.1016/s0076-6879(04)92005-8

Cited by 63 publications

(40 citation statements)

References 49 publications

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“…The significance of target structure has long been established for antisense oligonucleotides and trans-cleaving ribozymes (Zhao and Lemke 1998;Vickers et al 2000). For RNAi, however, this has been disputed in several reports, with one based on limited computational analysis (Reynolds et al 2004;Boese et al 2005).…”

Section: Introductionmentioning

confidence: 99%

Effect of target secondary structure on RNAi efficiency

Shao¹,

Chan²,

Maliyekkel³

et al. 2007

RNA

150

130

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RNA interference (RNAi) mediated by small interfering RNAs (siRNAs) or short hairpin RNAs (shRNAs) has become a powerful tool for gene knockdown studies. However, the levels of knockdown vary greatly. Here, we examine the effect of target disruption energy, a novel measure of target accessibility, along with other parameters that may affect RNAi efficiency. Based on target secondary structures predicted by the Sfold program, the target disruption energy represents the free energy cost for local alteration of the target structure to allow target binding by the siRNA guide strand. In analyses of 100 siRNAs and 101 shRNAs targeted to 103 endogenous human genes, we find that the disruption energy is an important determinant of RNAi activity and the asymmetry of siRNA duplex asymmetry is important for facilitating the assembly of the RNA-induced silencing complex (RISC). We estimate that target accessibility and duplex asymmetry can improve the target knockdown level significantly by nearly 40% and 26%, respectively. In the RNAi pathway, RISC assembly precedes target binding by the siRNA guide strand. Thus, our findings suggest that duplex asymmetry has significant upstream effect on RISC assembly and target accessibility has strong downstream effect on target recognition. The results of the analyses suggest criteria for improving the design of siRNAs and shRNAs.

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Section: Introductionmentioning

confidence: 99%

Effect of target secondary structure on RNAi efficiency

Shao¹,

Chan²,

Maliyekkel³

et al. 2007

RNA

150

130

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show abstract

“…The G/C content of siRNA molecules has been reported to be significant in siRNA-mediated silencing efficiency. It has been reported that efficient siRNA sequences contain 30-52% G/C content (35). Our siRNA-1 contained 38% G/C, siRNA-2 contained 44% G/C, and siRNA-3 contained 42% G/C, so that all three siRNAs were expected and shown to be effective at P-gp downregulation.…”

Section: Discussionmentioning

confidence: 69%

siRNA-Mediated Down-Regulation of P-glycoprotein in a Xenograft Tumor Model in NOD-SCID Mice

et al. 2011

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“…In light of this, many groups have devised algorithms to predict both the efficacy and the specificity of RNAi sequences. [31][32][33] Properties such as thermodynamic values, sequence asymmetry, and polymorphisms that contribute to RNA duplex stability are taken into account within these databases. 34 Artificial neural networks have been utilized to train algorithms based on the analysis of randomly selected siRNAs.…”

Section: Designing Potent Sirnasmentioning

confidence: 99%

From sequence to function: using RNAi to elucidate mechanisms of human disease

Wolters

MacKeigan

2008

Cell Death Differ

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RNA interference (RNAi) has emerged as one of the most powerful tools for functionally characterizing large sets of genomic data. Capabilities of RNAi place it at the forefront of high-throughput screens, which are able to span the human genome in search of novel targets. Although RNAi screens have been used to elucidate pathway components and discover potential drug targets in lower organisms, including Caenorhabditis elegans and Drosophila, only recently has the technology been advanced to a state in which large-scale screens can be performed in mammalian cells. In this review, we will evaluate the major advancements in the field of mammalian RNAi, specifically in terms of high-throughput assays. Crucial points of experimental design will be highlighted, as well as suggestions as to how to interpret and follow-up on potential cell death targets. Finally, we assess the prospective applications of high-throughput screens, the data they are capable of generating, and the potential for this technique to further our understanding of human disease.

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Mechanistic Insights Aid Computational Short Interfering RNA Design

Cited by 63 publications

References 49 publications

Effect of target secondary structure on RNAi efficiency

Effect of target secondary structure on RNAi efficiency

siRNA-Mediated Down-Regulation of P-glycoprotein in a Xenograft Tumor Model in NOD-SCID Mice

From sequence to function: using RNAi to elucidate mechanisms of human disease

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